RESUMO
BACKGROUND: Electronic visits (e-visits) are billable, asynchronous patient-initiated messages that require at least five minutes of medical decision-making by a provider. Unequal use of patient portal tools like e-visits by certain patient populations may worsen health disparities. To date, no study has attempted to qualitatively assess perceptions of e-visits in older adults. OBJECTIVE: In this qualitative study, we aimed to understand patient perceptions of e-visits, including their perceived utility, barriers to use, and care implications, with a focus on vulnerable patient groups. METHODS: We conducted a qualitative study using in-depth structured individual interviews with patients from diverse backgrounds to assess their knowledge and perceptions surrounding e-visits as compared with unbilled portal messages and other visit types. We used content analysis to analyze interview data. RESULTS: We conducted 20 interviews, all in adults older than 65 years. We identified 4 overarching coding categories or themes. First, participants were generally accepting of the concept of e-visits and willing to try them. Second, nearly two-thirds of the participants voiced a preference for synchronous communication. Third, participants had specific concerns about the name "e-visit" and when to choose this type of visit in the patient portal. Fourth, some participants indicated discomfort using or accessing technology for e-visits. Financial barriers to the use of e-visits was not a common theme. CONCLUSIONS: Our findings suggest that older adults are generally accepting of the concept of e-visits, but uptake may be limited due to their preference for synchronous communication. We identified several opportunities to improve e-visit implementation.
RESUMO
Antibody heavy chain (HC) and light chain (LC) variable region exons are assembled by V(D)J recombination. V(D)J junctional regions encode complementarity-determining-region 3 (CDR3), an antigen-contact region immensely diversified through nontemplated nucleotide additions ("N-regions") by terminal deoxynucleotidyl transferase (TdT). HIV-1 vaccine strategies seek to elicit human HIV-1 broadly neutralizing antibodies (bnAbs), such as the potent CD4-binding site VRC01-class bnAbs. Mice with primary B cells that express receptors (BCRs) representing bnAb precursors are used as vaccination models. VRC01-class bnAbs uniformly use human HC VH1-2 and commonly use human LCs Vκ3-20 or Vκ1-33 associated with an exceptionally short 5-amino-acid (5-aa) CDR3. Prior VRC01-class models had nonphysiological precursor levels and/or limited precursor diversity. Here, we describe VRC01-class rearranging mice that generate more physiological primary VRC01-class BCR repertoires via rearrangement of VH1-2, as well as Vκ1-33 and/or Vκ3-20 in association with diverse CDR3s. Human-like TdT expression in mouse precursor B cells increased LC CDR3 length and diversity and also promoted the generation of shorter LC CDR3s via N-region suppression of dominant microhomology-mediated Vκ-to-Jκ joins. Priming immunization with eOD-GT8 60mer, which strongly engages VRC01 precursors, induced robust VRC01-class germinal center B cell responses. Vκ3-20-based responses were enhanced by N-region addition, which generates Vκ3-20-to-Jκ junctional sequence combinations that encode VRC01-class 5-aa CDR3s with a critical E residue. VRC01-class-rearranging models should facilitate further evaluation of VRC01-class prime and boost immunogens. These new VRC01-class mouse models establish a prototype for the generation of vaccine-testing mouse models for other HIV-1 bnAb lineages that employ different HC or LC Vs.
Assuntos
Infecções por HIV , Soropositividade para HIV , HIV-1 , Vacinas , Camundongos , Humanos , Animais , Anticorpos Amplamente Neutralizantes , Anticorpos Neutralizantes , HIV-1/genética , Anticorpos Anti-HIV , DNA Nucleotidilexotransferase , Regiões Determinantes de Complementaridade/genética , Infecções por HIV/prevenção & controleRESUMO
SARS-CoV-2 Omicron subvariants have generated a worldwide health crisis due to resistance to most approved SARS-CoV-2 neutralizing antibodies and evasion of vaccination-induced antibodies. To manage Omicron subvariants and prepare for new ones, additional means of isolating broad and potent humanized SARS-CoV-2 neutralizing antibodies are desirable. Here, we describe a mouse model in which the primary B cell receptor (BCR) repertoire is generated solely through V(D)J recombination of a human VH1-2 heavy chain (HC) and, substantially, a human Vκ1-33 light chain (LC). Thus, primary humanized BCR repertoire diversity in these mice derives from immensely diverse HC and LC antigen-contact CDR3 sequences generated by nontemplated junctional modifications during V(D)J recombination. Immunizing this mouse model with SARS-CoV-2 (Wuhan-Hu-1) spike protein immunogens elicited several VH1-2/Vκ1-33-based neutralizing antibodies that bound RBD in a different mode from each other and from those of many prior patient-derived VH1-2-based neutralizing antibodies. Of these, SP1-77 potently and broadly neutralized all SARS-CoV-2 variants through BA.5. Cryo-EM studies revealed that SP1-77 bound RBD away from the receptor-binding motif via a CDR3-dominated recognition mode. Lattice light-sheet microscopy-based studies showed that SP1-77 did not block ACE2-mediated viral attachment or endocytosis but rather blocked viral-host membrane fusion. The broad and potent SP1-77 neutralization activity and nontraditional mechanism of action suggest that it might have therapeutic potential. Likewise, the SP1-77 binding epitope may inform vaccine strategies. Last, the type of humanized mouse models that we have described may contribute to identifying therapeutic antibodies against future SARS-CoV-2 variants and other pathogens.
Assuntos
COVID-19 , SARS-CoV-2 , Humanos , Camundongos , Animais , Glicoproteína da Espícula de Coronavírus/genética , Enzima de Conversão de Angiotensina 2 , Fusão de Membrana , Anticorpos Antivirais , Anticorpos Neutralizantes , Epitopos , Receptores de Antígenos de Linfócitos BRESUMO
IgH class switch recombination (CSR) replaces Cµ constant region (CH) exons with one of six downstream CHs by joining transcription-targeted double-strand breaks (DSBs) in the Cµ switch (S) region to DSBs in a downstream S region. Chromatin loop extrusion underlies fundamental CSR mechanisms including 3'IgH regulatory region (3'IgHRR)-mediated S region transcription, CSR center formation, and deletional CSR joining. There are 10 consecutive CTCF-binding elements (CBEs) downstream of the 3'IgHRR, termed the "3'IgH CBEs." Prior studies showed that deletion of eight 3'IgH CBEs did not detectably affect CSR. Here, we report that deletion of all 3'IgH CBEs impacts, to varying degrees, germline transcription and CSR of upstream S regions, except that of Sγ1. Moreover, deletion of all 3'IgH CBEs rendered the 6-kb region just downstream highly transcribed and caused sequences within to be aligned with Sµ, broken, and joined to form aberrant CSR rearrangements. These findings implicate the 3'IgH CBEs as critical insulators for focusing loop extrusion-mediated 3'IgHRR transcriptional and CSR activities on upstream CH locus targets.
Assuntos
Fator de Ligação a CCCTC/genética , Switching de Imunoglobulina/genética , Cadeias Pesadas de Imunoglobulinas/genética , Transcrição Gênica/imunologia , Animais , Anticorpos/genética , Anticorpos/imunologia , Linfócitos B/imunologia , Cromatina/genética , Cromatina/imunologia , Mutação em Linhagem Germinativa/genética , Switching de Imunoglobulina/imunologia , Camundongos , Sequências Reguladoras de Ácido Nucleico/genética , Sequências Reguladoras de Ácido Nucleico/imunologiaRESUMO
RAG endonuclease initiates Igh V(D)J recombination in progenitor B cells by binding a JH-recombination signal sequence (RSS) within a recombination centre (RC) and then linearly scanning upstream chromatin, presented by loop extrusion mediated by cohesin, for convergent D-RSSs1,2. The utilization of convergently oriented RSSs and cryptic RSSs is intrinsic to long-range RAG scanning3. Scanning of RAG from the DJH-RC-RSS to upstream convergent VH-RSSs is impeded by D-proximal CTCF-binding elements (CBEs)2-5. Primary progenitor B cells undergo a mechanistically undefined contraction of the VH locus that is proposed to provide distal VHs access to the DJH-RC6-9. Here we report that an inversion of the entire 2.4-Mb VH locus in mouse primary progenitor B cells abrogates rearrangement of both VH-RSSs and normally convergent cryptic RSSs, even though locus contraction still occurs. In addition, this inversion activated both the utilization of cryptic VH-RSSs that are normally in opposite orientation and RAG scanning beyond the VH locus through several convergent CBE domains to the telomere. Together, these findings imply that broad deregulation of CBE impediments in primary progenitor B cells promotes RAG scanning of the VH locus mediated by loop extrusion. We further found that the expression of wings apart-like protein homologue (WAPL)10, a cohesin-unloading factor, was low in primary progenitor B cells compared with v-Abl-transformed progenitor B cell lines that lacked contraction and RAG scanning of the VH locus. Correspondingly, depletion of WAPL in v-Abl-transformed lines activated both processes, further implicating loop extrusion in the locus contraction mechanism.
Assuntos
Linfócitos B/metabolismo , Proteínas de Ligação a DNA/metabolismo , Endonucleases/metabolismo , Proteínas de Homeodomínio/metabolismo , Cadeias Pesadas de Imunoglobulinas/genética , Conformação de Ácido Nucleico , Animais , Linfócitos B/citologia , Linfócitos B/enzimologia , Linhagem Celular , Células Cultivadas , Proteínas de Ligação a DNA/deficiência , Proteínas de Ligação a DNA/genética , Regulação para Baixo , Endonucleases/deficiência , Endonucleases/genética , Pontos de Checagem da Fase G1 do Ciclo Celular , Sequenciamento de Nucleotídeos em Larga Escala , Proteínas de Homeodomínio/genética , Camundongos , Camundongos Endogâmicos C57BL , Proteínas/genética , Proteínas/metabolismo , Recombinação V(D)J/genéticaRESUMO
We provide a protocol for generating forebrain structures in vivo from mouse embryonic stem cells (ESCs) via neural blastocyst complementation (NBC). We developed this protocol for studies of development and function of specific forebrain regions, including the cerebral cortex and hippocampus. We describe a complete workflow, from methods for modifying a given genomic locus in ESCs via CRISPR-Cas9-mediated editing to the generation of mouse chimeras with ESC-reconstituted forebrain regions that can be directly analyzed. The procedure begins with genetic editing of mouse ESCs via CRISPR-Cas9, which can be accomplished in ~4-8 weeks. We provide protocols to achieve fluorescent labeling of ESCs in ~2-3 weeks, which allows tracing of the injected, ESC-derived donor cells in chimeras generated via NBC. Once modified ESCs are ready, NBC chimeras are generated in ~3 weeks via injection of ESCs into genetically programmed blastocysts that are subsequently transferred into pseudo-pregnant fosters. Our in vivo brain organogenesis platform is efficient, allowing functional and systematic analysis of genes and other genomic factors in as little as 3 months, in the context of a whole organism.
Assuntos
Mapeamento Encefálico/métodos , Encéfalo/embriologia , Células-Tronco Embrionárias Murinas/fisiologia , Animais , Blastocisto , Diferenciação Celular , Quimera , Feminino , Masculino , Camundongos , Organogênese , FenótipoRESUMO
OBJECTIVE: To rapidly deploy a digital patient-facing self-triage and self-scheduling tool in a large academic health system to address the COVID-19 pandemic. MATERIALS AND METHODS: We created a patient portal-based COVID-19 self-triage and self-scheduling tool and made it available to all primary care patients at the University of California, San Francisco Health, a large academic health system. Asymptomatic patients were asked about exposure history and were then provided relevant information. Symptomatic patients were triaged into 1 of 4 categories-emergent, urgent, nonurgent, or self-care-and then connected with the appropriate level of care via direct scheduling or telephone hotline. RESULTS: This self-triage and self-scheduling tool was designed and implemented in under 2 weeks. During the first 16 days of use, it was completed 1129 times by 950 unique patients. Of completed sessions, 315 (28%) were by asymptomatic patients, and 814 (72%) were by symptomatic patients. Symptomatic patient triage dispositions were as follows: 193 emergent (24%), 193 urgent (24%), 99 nonurgent (12%), 329 self-care (40%). Sensitivity for detecting emergency-level care was 87.5% (95% CI 61.7-98.5%). DISCUSSION: This self-triage and self-scheduling tool has been widely used by patients and is being rapidly expanded to other populations and health systems. The tool has recommended emergency-level care with high sensitivity, and decreased triage time for patients with less severe illness. The data suggests it also prevents unnecessary triage messages, phone calls, and in-person visits. CONCLUSION: Patient self-triage tools integrated into electronic health record systems have the potential to greatly improve triage efficiency and prevent unnecessary visits during the COVID-19 pandemic.
Assuntos
Agendamento de Consultas , Betacoronavirus , Infecções por Coronavirus , Autoavaliação Diagnóstica , Sistemas Computadorizados de Registros Médicos , Pandemias , Participação do Paciente , Portais do Paciente , Pneumonia Viral , Triagem/métodos , Centros Médicos Acadêmicos , Adulto , COVID-19 , Infecções por Coronavirus/diagnóstico , Infecções por Coronavirus/epidemiologia , Humanos , Pneumonia Viral/diagnóstico , Pneumonia Viral/epidemiologia , SARS-CoV-2 , São Francisco , Autocuidado , Telemedicina/organização & administraçãoRESUMO
HIV-1 vaccine development aims to elicit broadly neutralizing antibodies (bnAbs) against diverse viral strains. In some HIV-1-infected individuals, bnAbs evolved from precursor antibodies through affinity maturation. To induce bnAbs, a vaccine must mediate a similar antibody maturation process. One way to test a vaccine is to immunize mouse models that express human bnAb precursors and assess whether the vaccine can convert precursor antibodies into bnAbs. A major problem with such mouse models is that bnAb expression often hinders B cell development. Such developmental blocks may be attributed to the unusual properties of bnAb variable regions, such as poly-reactivity and long antigen-binding loops, which are usually under negative selection during primary B cell development. To address this problem, we devised a method to circumvent such B cell developmental blocks by expressing bnAbs conditionally in mature B cells. We validated this method by expressing the unmutated common ancestor (UCA) of the human VRC26 bnAb in transgenic mice. Constitutive expression of the VRC26UCA led to developmental arrest of B cell progenitors in bone marrow; poly-reactivity of the VRC26UCA and poor pairing of the VRC26UCA heavy chain with the mouse surrogate light chain may contribute to this phenotype. The conditional expression strategy bypassed the impediment to VRC26UCA B cell development, enabling the expression of VRC26UCA in mature B cells. This approach should be generally applicable for expressing other bnAbs that are under negative selection during B cell development.
Assuntos
Vacinas contra a AIDS/imunologia , Anticorpos Anti-HIV/imunologia , Infecções por HIV/imunologia , Produtos do Gene env do Vírus da Imunodeficiência Humana/imunologia , Vacinas contra a AIDS/farmacologia , Animais , Anticorpos Neutralizantes/imunologia , Anticorpos Neutralizantes/fisiologia , Linfócitos B/imunologia , Linfócitos B/virologia , Modelos Animais de Doenças , Anticorpos Anti-HIV/farmacologia , Infecções por HIV/prevenção & controle , Infecções por HIV/virologia , Soropositividade para HIV/genética , Soropositividade para HIV/imunologia , HIV-1/efeitos dos fármacos , HIV-1/imunologia , HIV-1/patogenicidade , Humanos , Ativação Linfocitária/imunologia , Camundongos , Produtos do Gene env do Vírus da Imunodeficiência Humana/genéticaAssuntos
Vacinas contra a AIDS/genética , Afinidade de Anticorpos/genética , Linfócitos B/imunologia , Anticorpos Anti-HIV/imunologia , HIV-1/imunologia , Seleção Genética , Produtos do Gene env do Vírus da Imunodeficiência Humana/genética , Vacinas contra a AIDS/imunologia , Animais , Epitopos/genética , Técnicas de Introdução de Genes , Humanos , Macaca , Camundongos , Camundongos Mutantes , Mutação , Produtos do Gene env do Vírus da Imunodeficiência Humana/imunologiaRESUMO
BACKGROUND: The aim of this study was to analyze the impact of clinic nonattendance within the first year after kidney transplantation on graft-related outcomes. METHODS: Our retrospective analysis included all patients receiving their transplant (2007-2017) and receiving their long-term follow up at our center. Clinic nonattendance was extracted from electronic patient records and informatics systems, with highest clinic nonattenders stratified at the 75th percentile. RESULTS: Data were analyzed for 916 kidney allograft recipients, with median follow up 1168 days (interquartile range, 455-2073 days). Median number of missed transplant clinic visits in the first year was 5 (interquartile range, 3-7) and nonattenders were defined above the 75th percentile. Nonattenders versus attenders were more likely to be black, ABO-incompatible, repeat kidney transplant recipients but less likely to have pretransplantation diabetes. Nonattenders versus attenders had longer hospital stays after their transplant surgery in days (14.4 vs 12.2 respectively, P = 0.007), higher rate of delayed graft function (21.3% vs 12.8% respectively, P = 0.005), higher risk for 1-year rejection (12.5% vs 7.8% respectively, P = 0.044), worse 1-year estimated glomerular filtration rate in mL/min (47.0 vs 54.1, respectively, P = 0.002) and increased risk for death-censored graft loss by median follow (17.5% vs 12.0%, respectively, P = 0.013). In a Cox regression model, kidney transplant recipients defined as clinic nonattenders within the first postoperative year demonstrated a significantly increased rate of death-censored graft loss (hazard ratio, 1.983; 95% confidence interval, 1.061-3.707; P = 0.032). CONCLUSIONS: Kidney transplant recipients in the top quartile for nonattendance require additional support and supervision to help attenuate long-term risks to their graft function and survival.
RESUMO
Genetically modified mice are commonly generated by the microinjection of pluripotent embryonic stem (ES) cells into wild-type host blastocysts1, producing chimeric progeny that require breeding for germline transmission and homozygosity of modified alleles. As an alternative approach and to facilitate studies of the immune system, we previously developed RAG2-deficient blastocyst complementation2. Because RAG2-deficient mice cannot undergo V(D)J recombination, they do not develop B or T lineage cells beyond the progenitor stage2: injecting RAG2-sufficient donor ES cells into RAG2-deficient blastocysts generates somatic chimaeras in which all mature lymphocytes derive from donor ES cells. This enables analysis, in mature lymphocytes, of the functions of genes that are required more generally for mouse development3. Blastocyst complementation has been extended to pancreas organogenesis4, and used to generate several other tissues or organs5-10, but an equivalent approach for brain organogenesis has not yet been achieved. Here we describe neural blastocyst complementation (NBC), which can be used to study the development and function of specific forebrain regions. NBC involves targeted ablation, mediated by diphtheria toxin subunit A, of host-derived dorsal telencephalic progenitors during development. This ablation creates a vacant forebrain niche in host embryos that results in agenesis of the cerebral cortex and hippocampus. Injection of donor ES cells into blastocysts with forebrain-specific targeting of diphtheria toxin subunit A enables donor-derived dorsal telencephalic progenitors to populate the vacant niche in the host embryos, giving rise to neocortices and hippocampi that are morphologically and neurologically normal with respect to learning and memory formation. Moreover, doublecortin-deficient ES cells-generated via a CRISPR-Cas9 approach-produced NBC chimaeras that faithfully recapitulated the phenotype of conventional, germline doublecortin-deficient mice. We conclude that NBC is a rapid and efficient approach to generate complex mouse models for studying forebrain functions; this approach could more broadly facilitate organogenesis based on blastocyst complementation.
Assuntos
Blastocisto/citologia , Blastocisto/metabolismo , Organogênese , Prosencéfalo/citologia , Prosencéfalo/embriologia , Animais , Quimera/embriologia , Quimera/genética , Proteínas de Ligação a DNA/deficiência , Proteínas do Domínio Duplacortina , Feminino , Teste de Complementação Genética , Células Germinativas/metabolismo , Hipocampo/anatomia & histologia , Hipocampo/citologia , Hipocampo/embriologia , Hipocampo/fisiologia , Masculino , Camundongos , Camundongos Transgênicos , Proteínas Associadas aos Microtúbulos/deficiência , Células-Tronco Embrionárias Murinas/citologia , Células-Tronco Embrionárias Murinas/metabolismo , Neocórtex/anatomia & histologia , Neocórtex/citologia , Neocórtex/embriologia , Neocórtex/fisiologia , Neurônios/citologia , Neurônios/metabolismo , Neuropeptídeos/deficiência , Fenótipo , Prosencéfalo/anatomia & histologia , Prosencéfalo/fisiologiaRESUMO
OBJECTIVES: There are conflicting reports in the literature regarding outcomes after kidney transplant for patients of black ethnicity. To investigate further, we compared outcomes for black versus white kidney transplant recipients in a single UK transplant center. MATERIALS AND METHODS: We analyzed 1066 kidney transplant recipients (80 black patients, 986 white patients) within a single-center cohort (2007-2017) in the United Kingdom, with cumulative 4446 patient-year follow-up. Data were electronically extracted from the Department of Health Informatics database for every study recruit, with manual data linkage to the UK Transplant Registry (for graft survival, delayed graft function, and rejection data) and Office for National Statistics (for mortality data). Primary outcomes of interest were graft/patient survival. RESULTS: Black recipients have increased baseline risk profiles with longer wait times, difficulty in matching, worse HLA matching, more socioeconomic deprivation, and lower rates of living kidney donors. Postoperatively, black versus white recipients had increased risk for delayed graft function (34.3% vs 10.2%; P < .001), increased 1-year rejection (16.7% vs 7.3%; P = .012), higher 1-year creatinine levels (166 vs 138 mmol/L; P = .003), and longer posttransplant length of stay (14.5 vs 9.5 days; P = .020). Although black recipients did not have increased risk of death versus white recipients (10.0% vs 11.0%, respectively; P = .486), they did have increased risk for death-censored graft loss (23.8% vs 11.1%; P = .002). However, in an adjusted Cox regression model, black ethnicity was not associated with increased risk for death-censored graft loss (hazard ratio of 1.209, 95% confidence interval, 0.660-2.216; P = .539). CONCLUSIONS: Black kidney transplant recipients in the United Kingdom have increased risk of adverse graft-related outcomes due to high-risk baseline variables rather than their black ethnicity per se.
Assuntos
População Negra , Função Retardada do Enxerto/etnologia , Rejeição de Enxerto/etnologia , Sobrevivência de Enxerto , Transplante de Rim/efeitos adversos , População Branca , Adulto , Bases de Dados Factuais , Função Retardada do Enxerto/diagnóstico , Função Retardada do Enxerto/mortalidade , Feminino , Rejeição de Enxerto/diagnóstico , Rejeição de Enxerto/mortalidade , Disparidades nos Níveis de Saúde , Disparidades em Assistência à Saúde/etnologia , Humanos , Transplante de Rim/mortalidade , Masculino , Pessoa de Meia-Idade , Sistema de Registros , Estudos Retrospectivos , Fatores de Risco , Determinantes Sociais da Saúde/etnologia , Fatores de Tempo , Resultado do Tratamento , Reino Unido/epidemiologiaRESUMO
Thanatophoric dysplasia is a type of short-limbed neonatal dwarfism that is usually lethal in the perinatal period. It is characterized by short limbs, a narrow, bell-shaped thorax, macrocephaly with a prominent forehead, and flattened vertebral bodies. These malformations result from autosomal dominant mutations in the fibroblast growth factor receptor 3 (FGFR3) gene. In this report, we describe a novel FGFR3 insertion mutation in a fetus with shortened limbs, curved femurs, and a narrow thorax. The diagnosis of thanatophoric dysplasia type 1 was suspected clinically, and FGFR3 sequencing showed a c.742_743insTGT variant, which predicts p.R248delinsLC. In vivo studies in zebrafish demonstrated that this mutation resulted in the overexpression of zebrafish Fgfr3, leading to the over-activation of downstream signaling and dorsalized embryos. To date, no insertions or deletions in FGFR3 have been reported to cause thanatophoric dysplasia types 1 or 2; therefore, this represents the first report to describe such a mutation. © 2016 Wiley Periodicals, Inc.
Assuntos
Estudos de Associação Genética , Mutagênese Insercional , Fenótipo , Receptor Tipo 3 de Fator de Crescimento de Fibroblastos/deficiência , Receptor Tipo 3 de Fator de Crescimento de Fibroblastos/genética , Displasia Tanatofórica/diagnóstico , Displasia Tanatofórica/genética , Aborto Induzido , Alelos , Animais , Autopsia , Éxons , Feminino , Feto , Expressão Gênica , Genótipo , Humanos , Mutação , Gravidez , Peixe-ZebraRESUMO
Metabolic engineers strive to improve the production yields of microbial fermentations, sometimes by mutating the genomes of production strains. Some mutations are detrimental to the health of the organism, so a quantitative and mechanistic understanding of the trade-offs could inform better designs. We employed the bacterial luciferase operon (luxABCDE), which uses ubiquitous energetic cofactors (NADPH, ATP, FMNH2, acetyl-CoA) from the host cell, as a proxy for a novel anabolic pathway. The strains in the Escherichia coli Keio collection, each of which contains a single deletion of a non-essential gene, represent mutational choices that an engineer might make to optimize fermentation yields. The Keio strains and the parental BW25113 strain were transformed with a luxABCDE expression vector. Each transformant was propagated in defined M9 medium at 37 °C for 48 hours; the cell density (optical density at 600 nanometers, OD600) and luminescence were measured every 30 minutes. The trade-offs were visualized by plotting the maximum growth rate and luminescence/OD600 of each transformant across a "production possibility frontier". Our results show that some loss-of-function mutations enhance growth in vitro or light production, but that improvement in one trait generally comes at the expense of the other.
Assuntos
Escherichia coli/crescimento & desenvolvimento , Escherichia coli/genética , Metabolismo/genética , Metabolismo/fisiologia , Deleção de Sequência/genética , Contagem de Células , Proteínas de Escherichia coli/genética , Fermentação/genética , Fermentação/fisiologia , Luz , Luciferases Bacterianas/genética , Luciferases Bacterianas/metabolismo , Luminescência , Óperon/genéticaRESUMO
Medicaid physician fees increased 15.1 percent, on average, between 2003 and 2008. This was below the general rate of inflation, resulting in a reduction in real fees. Only primary care fees grew at the rate of inflation-20 percent between 2003 and 2008. However, because of slow growth in Medicare fees, Medicaid fees closed a small portion of their ongoing gap relative to Medicare-growing from 69 percent to 72 percent of Medicare. The increase in Medicaid fees relative to Medicare fees resulted from relative increases for primary care and obstetrical services, but not for other services.
Assuntos
Honorários Médicos/tendências , Medicaid/tendências , Adulto , Idoso , Administração de Caso/tendências , Tabela de Remuneração de Serviços/tendências , Planos de Pagamento por Serviço Prestado/tendências , Previsões , Sistemas Pré-Pagos de Saúde/tendências , Acessibilidade aos Serviços de Saúde/tendências , Humanos , Inflação/tendências , Cobertura do Seguro/tendências , Medicaid/economia , Medicare/economia , Medicare/tendências , Medicare Assignment/tendências , Atenção Primária à Saúde/tendências , Estados UnidosRESUMO
The "patient-centered medical home" has been promoted as an enhanced model of primary care. Based on a literature review and interviews with practicing physicians, we find that medical home advocates and physicians have somewhat different, although not necessarily inconsistent, expectations of what the medical home should accomplish-from greater responsiveness to the needs of all patients to increased focus on care management for patients with chronic conditions. As the medical home concept is further developed, it will be important to not overemphasize redesign of practices at the expense of patient-centered care, which is the hallmark of excellent primary care.
Assuntos
Assistência Centrada no Paciente/organização & administração , Administração da Prática Médica/organização & administração , Atenção Primária à Saúde/normas , Doença Crônica/terapia , Pesquisa sobre Serviços de Saúde , Humanos , Inovação Organizacional , Estados UnidosRESUMO
Tuberous sclerosis complex is an autosomal dominant neurocutaneous disorder marked by hamartoma growth in multiple organ systems. We performed mutational analyses on 325 individuals with definite tuberous sclerosis complex diagnostic status. We identified mutations in 72% (199/257) of de novo and 77% (53/68) of familial cases, with 17% of mutations in the TSC1 gene and 50% in the TSC2 gene. There were 4% unclassified variants and 29% with no mutation identified. Genotype/phenotype analyses of all observed tuberous sclerosis complex findings in probands were performed, including several clinical features not analyzed in two previous large studies. We showed that patients with TSC2 mutations have significantly more hypomelanotic macules and learning disability in contrast to those with TSC1 mutations, findings not noted in previous studies. We also observed results consistent with two similar studies suggesting that individuals with mutations in TSC2 have more severe symptoms. On performing meta-analyses of our data and the other two largest studies in the literature, we found significant correlations for several features that individual studies did not have sufficient power to conclude. Male patients showed more frequent neurologic and eye symptoms, renal cysts, and ungual fibromas. Correlating genotypes with phenotypes should facilitate the disease management of tuberous sclerosis complex.
Assuntos
Predisposição Genética para Doença/genética , Fenótipo , Esclerose Tuberosa/genética , Esclerose Tuberosa/patologia , Proteínas Supressoras de Tumor/genética , Adolescente , Adulto , Criança , Pré-Escolar , Análise Mutacional de DNA , Feminino , Genótipo , Humanos , Lactente , Masculino , Fatores Sexuais , Esclerose Tuberosa/complicações , Proteína 1 do Complexo Esclerose Tuberosa , Proteína 2 do Complexo Esclerose Tuberosa , Estados UnidosAssuntos
Doenças Renais Policísticas/complicações , Doenças Renais Policísticas/genética , Esclerose Tuberosa/genética , Substituição de Aminoácidos , Cromossomos Humanos Par 16 , Cromossomos Humanos Par 4 , Cromossomos Humanos Par 9 , Feminino , Deleção de Genes , Frequência do Gene , Genes Dominantes , Ligação Genética , Marcadores Genéticos , Humanos , Incidência , Masculino , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Mutação de Sentido Incorreto , Linhagem , Penetrância , Doenças Renais Policísticas/patologia , Proteínas/genética , Proteínas/metabolismo , Análise de Sequência de DNA , Índice de Gravidade de Doença , Canais de Cátion TRPP , Triptofano/metabolismo , Esclerose Tuberosa/epidemiologia , Esclerose Tuberosa/metabolismo , Esclerose Tuberosa/patologia , Proteína 1 do Complexo Esclerose Tuberosa , Proteína 2 do Complexo Esclerose Tuberosa , Proteínas Supressoras de Tumor/genética , Proteínas Supressoras de Tumor/metabolismoRESUMO
This study sought to identify if differences existed in risk comprehension and risk format understanding between genetic counseling patients of Hispanic and Caucasian ethnicity. A total of 107 questionnaires were collected, 56 from Hispanic patients, and 51 from Caucasian controls. Of the total population 41.1% (44/107) could not demonstrate sufficient risk understanding, which was 71.4% (40/56) of Hispanics and 7.8% (4/51) of Caucasians. Fractions were the best-understood format for all participants. However, both Hispanics and Caucasians had difficulties with the percentage risk format. Discrepancies were also noted in qualitative word format understanding. Awareness of differences in risk comprehension may affect the selection of counseling techniques and strategies utilized by genetic counselors when educating patients about risk related information.
Assuntos
Atitude Frente a Saúde/etnologia , Aconselhamento Genético , Hispânico ou Latino/psicologia , Comparação Transcultural , Diversidade Cultural , Tomada de Decisões , Demografia , Humanos , Fatores de Risco , Inquéritos e Questionários , População BrancaRESUMO
Tuberous sclerosis is an autosomal dominant human disorder caused by inactivating mutations to either the TSC1 or TSC2 tumour suppressor gene. Hamartin and tuberin, the TSC1 and TSC2 gene products, interact and the tuberin-hamartin complex inhibits cell growth by antagonising signal transduction to downstream effectors of the mammalian target of rapamycin (mTOR) through the small GTPase rheb. Previously, we showed that pathogenic tuberin amino-acid substitutions disrupt the tuberin-hamartin complex. Here, we investigate how these mutations affect the role of tuberin in the control of signal transduction through mTOR. Our data indicate that specific amino-acid substitutions have distinct effects on tuberin function.