Evolutionary trajectories of small cell lung cancer under therapy.

The evolutionary processes that underlie the marked sensitivity of small cell lung cancer (SCLC) to chemotherapy and rapid relapse are unknown1-3. Here we determined tumour phylogenies at diagnosis and throughout chemotherapy and immunotherapy by multiregion sequencing of 160 tumours from 65 patients. Treatment-naive SCLC exhibited clonal homogeneity at distinct tumour sites, whereas first-line platinum-based chemotherapy led to a burst in genomic intratumour heterogeneity and spatial clonal diversity. We observed branched evolution and a shift to ancestral clones underlying tumour relapse. Effective radio- or immunotherapy induced a re-expansion of founder clones with acquired genomic damage from first-line chemotherapy. Whereas TP53 and RB1 alterations were exclusively part of the common ancestor, MYC family amplifications were frequently not constituents of the founder clone. At relapse, emerging subclonal mutations affected key genes associated with SCLC biology, and tumours harbouring clonal CREBBP/EP300 alterations underwent genome duplications. Gene-damaging TP53 alterations and co-alterations of TP53 missense mutations with TP73, CREBBP/EP300 or FMN2 were significantly associated with shorter disease relapse following chemotherapy. In summary, we uncover key processes of the genomic evolution of SCLC under therapy, identify the common ancestor as the source of clonal diversity at relapse and show central genomic patterns associated with sensitivity and resistance to chemotherapy.

Interprofessional collaboration and patient-reported outcomes: a secondary data analysis based on large scale survey data.

BACKGROUND: While interprofessional collaboration (IPC) is widely considered a key element of comprehensive patient treatment, evidence focusing on its impact on patient-reported outcomes (PROs) is inconclusive. The aim of this study was to investigate the association between employee-rated IPC and PROs in a clinical inpatient setting. METHODS: We conducted a secondary data analysis of the entire patient and employee reported data collected by the Picker Institute Germany in cross-sectional surveys between 2003 and 2016. Individual patient data from departments within hospitals was matched with employee survey data from within 2 years of treatment at the department-level. Items assessing employee-rated IPC (independent variables) were included in Principal Component Analysis (PCA). All questions assessing PROs (overall satisfaction, less discomforts, complications, treatment success, willingness to recommend) served as main dependent variables in ordered logistic regression analyses. Results were adjusted for multiple hypothesis testing as well as patients' and employees' gender, age, and education. RESULTS: The data set resulted in 6154 patients from 19 hospitals respective 103 unique departments. The PCA revealed three principal components (department-specific IPC, interprofessional organization, and overall IPC), explaining 67% of the total variance. The KMO measure of sampling adequacy was .830 and Bartlett's test of sphericity highly significant (p < 0.001). An increase of 1 SD in department-specific IPC was associated with a statistically significant chance of a higher (i.e., better) PRO-rating about complications after discharge (OR 1.07, 95% CI 1.00-1.13, p = 0.029). However, no further associations were found. Exploratory analyses revealed positive coefficients of department-specific IPC on all PROs for patients which were treated in surgical or internal medicine departments, whereas results were ambiguous for pediatric patients. CONCLUSIONS: The association between department-level IPC and patient-level PROs remains - as documented in previous literature - unclear and results are of marginal effect sizes. Future studies should keep in mind the different types of IPC, their specific characteristics and possible effect mechanisms. TRIAL REGISTRATION: Study registration: Open Science Framework (DOI https://doi.org/10.17605/OSF.IO/2NYAX ); Date of registration: 09 November 2021.

Interprofessional collaboration and patient-reported outcomes in inpatient care: a systematic review.

BACKGROUND: Interprofessional collaboration (IPC) is seen as the "gold standard" of comprehensive care, but credible evidence concerning the effects on patient-reported outcomes (PRO) is lacking. The aim of this systematic review is to study the effect of IPC on PRO in inpatient care. METHODS: We systematically searched six electronic databases (PubMed, Web of Science/Social Science Citation Index, CENTRAL (Cochrane Library), Current Contents (LIVIVO), CINAHL, and Embase) for studies published between 1997 and 2021. Additional studies were identified through citation tracking, manually searching the Internet and Google Scholar, and consultation of experts. Risk of bias (RoB) was assessed using the RoB 2 tool for randomized controlled trials (RCTs) and ROBINS-I for non-randomized studies (NRS). The included controlled before-and-after study (CBA) was assessed using both the ROBINS-I and the Effective Practice and Organization of Care (EPOC) quality criteria. Results were synthesized through narrative description, grouping, and thematic analysis of extracted data. RESULTS: The search yielded 10,213 records, from which 22 studies (16 RCTs, five NRS, and one CBA) fulfilled the inclusion criteria. In all but five studies, RoB was assessed as being high (RoB 2) resp. critical or serious (ROBINS-I). Within these 22 studies, nine inductively derived outcomes were assessed: (i) quality of life, (ii) coping, (iii) functional ability and health status, (iv) psychiatric morbidity, (v) pain, (vi) managing one's own health care, (vii) treatment success, (viii) satisfaction, and (ix) therapeutic relationship. While some studies do not report effect estimates, and some of the reported effects appear to be imprecisely estimated, the overall results indicate that IPC may affect PRO positively across all outcomes. CONCLUSIONS: Due to high clinical heterogeneity and high RoB, the question whether IPC affects PRO cannot be answered conclusively. Methodically rigorous studies are needed in order to answer the question of effectiveness of IPC. SYSTEMATIC REVIEW REGISTRATION: PROSPERO CRD42017073900.

Music Listening as a Postanesthesia Care Unit (PACU) Nursing Intervention for Laparoscopic Radical Prostatectomy Patients: A Randomized Comparative Clinical Trial.

PURPOSE: Randomized comparative mixed method approach with qualitative inquiry study's aim sought to determine if there was a difference in pre/post-intervention State-Trait Anxiety Inventory (STAI) scores and postanesthesia pain scores between two music listening groups of laparoscopic radical prostatectomy patients. DESIGN: Prospective randomized comparative mixed method approach with a qualitative inquiry. METHODS: Sample size of 77 male participants assigned by a table of random numbers to Spotify patient-preferred music selection Group I (n = 37) or minimalist hypnotic music with guided relaxation breathing (MHMGRB) instructional narrative Group II (n = 40). Outcome measures used patients' STAI questionnaire and reported PACU admission and discharge pain scores. RESULTS: Both Groups I and II had reduced pain scores at discharge as compared to admission; both groups had a significant reduction (P = .046 Group I, and Group II (P = .002), but changes for comparative groups (I and II) were not significant between the two groups (P = .53). CONCLUSIONS: Study revealed that both patient-preferred selected music and MHMGRB can meaningfully reduce patients' anxiety and PACU pain scores.

Pressure Injury Prevention for Complex Cardiovascular Patients in the Operating Room and Intensive Care Unit: A Quality Improvement Project.

PURPOSE: The purpose of our project was to examine the effect of an alternating pressure (AP) overlay on hospital-acquired pressure injury (HAPI) in high-risk cardiovascular surgical patients. PARTICIPANTS AND SETTING: This quality improvement (QI) initiative was conducted in a core group of 8 cardiovascular operating room (OR) suites and 1 cardiovascular surgical critical intensive care unit (ICU) in a large Indiana-based academic hospital. The sample comprised adult patients who underwent complex cardiovascular surgical procedures and those in the cardiovascular surgical ICU with extracorporeal membrane oxygenation (ECMO), ventricular assistive device (VAD), and undergoing heart and/or lung transplant, or open chest procedures. APPROACH: The AP overlay was placed on OR cardiovascular foam surfaces and on selected ICU support surfaces for patients who met inclusion criteria. We used a pre/postcomparative QI design to assess outcomes including OR-related HAPI rates, ICU aggregate unit HAPI data, related costs, and staff satisfaction during the 3-month project period. OUTCOMES: Operating room-related HAPIs were reduced from 8/71 (11%) preintervention to 0/147 (0%) postintervention (P = .008), resulting in a cost avoidance of $323,048 and positive staff satisfaction (mean = 3.85; 1- to 4-point Likert scale). No adverse outcomes occurred. Although not significant, ICU HAPI rates decreased from 10 to 7 pre/postintervention (P = .29), demonstrating a 14% HAPI reduction with a cost avoidance of $121,143. The ICU incidence density decreased from 3.57 to 3.24; however, there was no decrease in ICU monthly unit prevalence. Critical care staff satisfaction was positive (mean = 2.95; 1- to 4-point Likert scale) with most staff members preferring the AP overlay to a fluid immersion surface. A cost savings of 48% (AP overlay vs fluid immersion rental) was identified in the ICU. IMPLICATIONS FOR PRACTICE: We achieved fewer HAPIs and reduced costs and observed positive staff satisfaction, along with no adverse events with the use of the AP overlay. Further research is needed to determine the safety and efficacy of this device for this pressure injury prevention option for immobile patients in both the OR and the ICU.

[GRADE guidelines 20: Assessing the certainty of evidence in the importance of outcomes or values and preferences - inconsistency, imprecision, and other domains]. / GRADE Leitlinien: 20. Untersuchung der Vertrauenswürdigkeit in die Evidenz zur Beurteilung der Wichtigkeit von Endpunkten oder Werten und Präferenzen ­ Inkonsistenz, unzureichende Präzision und andere Domänen.

OBJECTIVE: To provide Grading of Recommendations, Assessment, Development, and Evaluation (GRADE) guidance for assessing inconsistency, imprecision, and other domains for the certainty of evidence about the relative importance of outcomes. STUDY DESIGN AND SETTING: We applied the GRADE domains to rate the certainty of evidence in the importance of outcomes to several systematic reviews, iteratively reviewed draft guidance, and consulted GRADE members and other stakeholders for feedback. RESULTS: We describe the rationale for considering the remaining GRADE domains when rating the certainty in a body of evidence for the relative importance of outcomes. As meta-analyses are not common in this context, inconsistency and imprecision assessments are challenging. Furthermore, confusion exists about inconsistency, imprecision, and true variability in the relative importance of outcomes. To clarify this issue, we suggest that the true variability is neither equivalent to inconsistency nor imprecision. Specifically, inconsistency arises from population, intervention, comparison and outcome and methodological elements that should be explored and, if possible, explained. The width of the confidence interval and sample size inform judgments about imprecision. We also provide suggestions on how to detect publication bias and discuss the domains to rate up the certainty. CONCLUSION: We provide guidance and examples for rating inconsistency, imprecision, and other domains for a body of evidence describing the relative importance of outcomes.

Mechanochemical control of epidermal stem cell divisions by B-plexins.

The precise spatiotemporal control of cell proliferation is key to the morphogenesis of epithelial tissues. Epithelial cell divisions lead to tissue crowding and local changes in force distribution, which in turn suppress the rate of cell divisions. However, the molecular mechanisms underlying this mechanical feedback are largely unclear. Here, we identify a critical requirement of B-plexin transmembrane receptors in the response to crowding-induced mechanical forces during embryonic skin development. Epidermal stem cells lacking B-plexins fail to sense mechanical compression, resulting in disinhibition of the transcriptional coactivator YAP, hyperproliferation, and tissue overgrowth. Mechanistically, we show that B-plexins mediate mechanoresponses to crowding through stabilization of adhesive cell junctions and lowering of cortical stiffness. Finally, we provide evidence that the B-plexin-dependent mechanochemical feedback is also pathophysiologically relevant to limit tumor growth in basal cell carcinoma, the most common type of skin cancer. Our data define a central role of B-plexins in mechanosensation to couple cell density and cell division in development and disease.

[GRADE Guidelines: 19. Assessing the certainty of evidence in the importance of outcomes or values and preferences: Risk of bias and indirectness]. / GRADE-Leitlinien: 19. Bewertung der Vertrauenswürdigkeit der Evidenz für die Bedeutung von Endpunkten oder Werten und Präferenzen ­ Risiko für Bias und Indirektheit.

OBJECTIVES: The Grading of Recommendations Assessment, Development, and Evaluation (GRADE) working group defines patient values and preferences as the relative importance patients place on the main health outcomes. We provide GRADE guidance for assessing the risk of bias and indirectness domains for certainty of evidence about the relative importance of outcomes. STUDY DESIGN AND SETTING: We applied the GRADE domains to rate the certainty of evidence in the importance of outcomes to several systematic reviews, iteratively reviewed draft guidance and consulted GRADE members and other stakeholders for feedback. RESULTS: This is the first of two articles. A body of evidence addressing the importance of outcomes starts at "high certainty"; concerns with risk of bias, indirectness, inconsistency, imprecision, and publication bias lead to downgrading to moderate, low, or very low certainty. We propose the following subdomains of risk of bias: selection of the study population, missing data, the type of measurement instrument, and confounding; we have developed items for each subdomain. The population, intervention, comparison, and outcome elements associated with the evidence determine the degree of indirectness. CONCLUSION: This article provides guidance and examples for rating the risk of bias and indirectness for a body of evidence summarizing the importance of outcomes.

Neurological repercussions of changes in cerebral blood flow in neonates undergoing cardiovascular surgery.

INTRODUCTION: Surgery for congenital heart disease can generate cerebral perfusion-associated alterations with neurological repercussions. OBJECTIVE: To analyze the relationship of peri-surgical cerebrovascular resistance index (RI) with mediate neurological functions after congenital heart disease surgery. METHOD: Prospective cohort study of 34 neonates in whom basilar artery RI, serum oxygen, carbon dioxide and lactate levels were determined before and after palliative or corrective procedures. We related pre-surgical RI with post-surgical ability to initiate the enteral route or to restore unassisted spontaneous breathing. RESULTS: Three groups were formed: 79 neonates with high RI (> 0.73), 73 with normal RI (0.63 to 0.73) and eight with low RI (< 0.63). In the former group, high RI persisted in the postoperative period, with persistent hyperlactatemia and hypoxia; in 86 %, the enteral route could not be initiated, and neither could assisted ventilation be withdrawn. In the second group, RI remained within normal values. In the third group, although RI, serum lactate and arterial oxygen pressure tended to normalize, 71 % had severe neurological damage. CONCLUSIONS: RI changes were common, although neurological damage appears to occur more commonly when RI remains high, possibly associated with low cerebral blood flow.

INTRODUCCIÓN: La cirugía de cardiopatías congénitas puede generar alteraciones perfusorias cerebrales con repercusión neurológica. OBJETIVO: Analizar la relación del índice de resistencia (IR) vascular cerebral periquirúrgico con funciones neurológicas mediatas posteriores a cirugía de cardiopatía congénita. MÉTODO: Estudio de cohorte prospectivo de 34 neonatos en quienes se determinó IR de la arteria basilar, niveles séricos de oxígeno, dióxido de carbono y lactato, antes y después de procedimientos paliativos o correctivos. Relacionamos el IR prequirúrgico con la capacidad posquirúrgica para iniciar la vía enteral o restablecer la respiración espontánea no asistida. RESULTADOS: Se integraron tres grupos: 79 neonatos con IR alto > 0.73, 73 con IR normal de 0.63 a 0.73 y ocho con IR bajo < 0.63. En los primeros persistió IR elevado en el posquirúrgico, con hiperlactatemia e hipoxia persistentes; en 86 % no se logró iniciar la vía enteral ni retirar la ventilación asistida. En los segundos, el IR se mantuvo en valores normales. En los terceros, si bien el IR, el lactato sérico y la presión arterial de oxígeno tendieron a normalizarse, 71 % presentó daño neurológico grave. CONCLUSIONES: Los cambios en el IR fueron frecuentes, aunque el daño neurológico parece presentarse más cuando el IR se mantiene alto, posiblemente asociado con flujos cerebrales bajos.

Repercusión neurológica por cambios en el flujo sanguíneo cerebral en neonatos sometidos a cirugía cardiovascular / Neurological repercussions of changes in cerebral blood flow in neonates undergoing cardiovascular surgery

Resumen Introducción: La cirugía de cardiopatías congénitas puede generar alteraciones perfusorias cerebrales con repercusión neurológica. Objetivo: Analizar la relación del índice de resistencia (IR) vascular cerebral periquirúrgico con funciones neurológicas mediatas posteriores a cirugía de cardiopatía congénita. Método: Estudio de cohorte prospectivo de 34 neonatos en quienes se determinó IR de la arteria basilar, niveles séricos de oxígeno, dióxido de carbono y lactato, antes y después de procedimientos paliativos o correctivos. Relacionamos el IR prequirúrgico con la capacidad posquirúrgica para iniciar la vía enteral o restablecer la respiración espontánea no asistida. Resultados: Se integraron tres grupos: 17 neonatos con IR alto > 0.73, cinco con IR normal de 0.63 a 0.73 y seis con IR bajo < 0.63. En los primeros persistió IR alto en el posquirúrgico, con hiperlactatemia e hipoxia persistentes; en 86 % no se logró iniciar la vía enteral ni retirar la ventilación asistida. En los segundos, el IR se mantuvo en valores normales. En los terceros, si bien el IR, el lactato sérico y la presión arterial de oxígeno tendieron a normalizarse, 71 % presentó daño neurológico grave. Conclusiones: Los cambios en el IR fueron frecuentes, aunque el daño neurológico parece presentarse más cuando el IR se mantiene alto, posiblemente asociado a flujos cerebrales bajos.

Abstract Introduction: Surgery for congenital heart disease can generate cerebral perfusion-associated alterations with neurological repercussions. Objective: To analyze the relationship of peri-surgical cerebrovascular resistance index (RI) with mediate neurological functions after congenital heart disease surgery. Method: Prospective cohort study of 34 neonates in whom basilar artery RI, serum oxygen, carbon dioxide and lactate levels were determined before and after palliative or corrective procedures. We related pre-surgical RI with post-surgical ability to initiate the enteral route or to restore unassisted spontaneous breathing. Results: Three groups were formed: 17 neonates with high RI (> 0.73), five with normal RI (0.63-0.73) and six with low RI (< 0.63). In the former group, high RI persisted in the postoperative period, with persistent hyperlactatemia and hypoxia; in 86%, the enteral route could not be initiated, and neither could assisted ventilation be withdrawn. In the second group, IR remained within normal values. In the third group, although RI, serum lactate and arterial oxygen pressure tended to normalize, 71% had severe neurological damage. Conclusions: RI changes were common, although neurological damage appears to occur more commonly when RI remains high, possibly associated with low cerebral blood flow.

[GRADE: Evidence to Decision (EtD) frameworks - a systematic and transparent approach to making well informed healthcare choices. 2: Clinical guidelines]. / GRADE: Von der Evidenz zur Empfehlung oder Entscheidung - ein systematischer und transparenter Ansatz, um gut informierte Entscheidungen im Gesundheitswesen zu treffen. 2: Klinische Praxisleitlinien.

Clinical guidelines are based on the best available evidence and produced systematically. In this context, the transparent presentation of the decision-making process from evidence to recommendation is indispensable. The Evidence to Decision (EtD) frameworks for clinical practice recommendations enables guideline panels to structure their approach and make it comprehensible. The EtD frameworks include three main sections: formulating the question, assessing the evidence and "Additional considerations" for each criterion, and drawing conclusions. This article focuses on the selection and operationalization of those criteria of the EtD framework that are relevant for clinical recommendations in guidelines. These include the priority of the problem, benefits and harms, certainty of the evidence, importance of the outcome, balance, resource use, equity, acceptability and feasibility. To make a recommendation, a panel must consider the implication and importance of each of the above judgments. The EtD framework helps ensure consideration of key criteria that determine whether an intervention should be recommended and that judgments are informed by the best available evidence.

International comparison of minimum volume standards for hospitals.

INTRODUCTION: Minimum volume standards have been implemented in various countries for quality or safety policies. We present minimum volume standards in an international comparison, focusing on regulatory approaches, selected sets of procedures and thresholds as well as predetermined consequences of non-compliance. MATERIALS AND METHODS: We combined a comprehensive literature search in electronic databases in March 2016 with a hand-search of governmental and related organisations' webpages. We also contacted international experts to verify the information we found in the literature and to obtain additional data. RESULTS: Minimum volume standards have been introduced in different countries predominantly for highly specialized surgical procedures. The same evidence has led to different definitions and ways of implementation of minimum volume standards in Germany, Canada (Ontario), the Netherlands, Switzerland, and Austria. The regulatory approaches to minimum volume standards and the predetermined consequences of non-compliance differ across the countries. CONCLUSION: The sets of procedures for which minimum volume standards and corresponding thresholds have been introduced vary across countries, possibly due to different regulatory approaches. In addition, key attributes of the health care system might affect the development and implementation of minimum volume standards. Therefore, it is not feasible to formulate uniform recommendations that are applicable to all countries. Our results provide a comprehensive overview of international minimum volume standards and can be used to inform policy decisions.

Interprofessional collaboration and patient-reported outcomes in inpatient care: protocol for a systematic review.

BACKGROUND: Interprofessional collaboration (IPC) is a core demand of policymakers, funding parties, and health care professionals in practice. Although the causal mechanism from increased IPC to improved patient outcomes seems to be intuitive, there is a lack of credible causal evidence concerning the effects not only on the objective but also on the subjective patient outcomes. The aim of the planned systematic review is to focus on the effect of IPC on patient-reported outcomes and experiences in inpatient care. METHODS: A systematic literature review will be undertaken by searching the following electronic databases: PubMed, Web of Science/Social Science Citation Index, Cochrane Library (CENTRAL), Current Contents (LIVIVO), CINAHL, and EMBASE. Additional studies will be identified through forward and backward citation tracking, manually searching the Internet and Google Scholar, and consultation of experts. Data will be synthesized through narrative description, grouping, and thematic analysis of the extracted data. If heterogeneity for some studies and outcomes is sufficiently low, a quantitative meta-analysis of effect sizes and standard errors will be applied. DISCUSSION: The systematic review will synthesize the evidence regarding the effectiveness of IPC and how it is perceived by patients in inpatient care. As the patients' perspective becomes increasingly relevant in the context of quality improvement, the results can help decision-makers in policy- and health care institutions to understand and develop strategies to ensure a high quality of care. SYSTEMATIC REVIEW REGISTRATION: PROSPERO registration number: CRD42017073900 ; date of registration in PROSPERO 07 August 2017.

Adaption and Validation of the Picker Employee Questionnaire With Hospital Midwives.

OBJECTIVE: To describe the adaption and psychometric testing of the Picker Employee Questionnaire to measure work environment, work experience, and employee engagement with midwives. DESIGN: Expert interviews, cognitive testing, and online survey for data collection. SETTING: Obstetric departments in Germany. PARTICIPANTS: Midwives employed in German obstetric departments: 3,867 were invited to take part, and 1,692 (44%) responded to the survey. METHODS: Questionnaire adaption involved expert interviews and cognitive testing. Psychometric evaluation was done via exploratory factor analysis, reliability analysis, and construct validity assessment. RESULTS: The adaption of the Picker Employee Questionnaire resulted in a tool with 75 closed questions referring to central aspects of work environment, experience, and engagement. Factor analysis yielded 10 factors explaining 51% of the variance. Themes covered were Support from Management (Immediate Superior and Hospital Management), Workload, Overtime, Scheduling, Education and Training, Interaction with Colleagues (Midwives, Physicians, and Nurses), and Engagement. Eight scales had a Cronbach's alpha coefficient of 0.7 or greater; the remaining two were 0.6 or less. The questionnaire distinguished between different subgroups of midwives and hospitals. CONCLUSION: The questionnaire is well suited for the measurement of midwives' work experience, environment, and engagement. It is a useful tool that supports employers and human resource managers in shaping and motivating an efficient work environment for midwives.

Diversifying selection and color-biased dispersal in the asp viper.

BACKGROUND: The presence of intraspecific color polymorphism can have multiple impacts on the ecology of a species; as a consequence, particular color morphs may be strongly selected for in a given habitat type. For example, the asp viper (Vipera aspis) shows a high level of color polymorphism. A blotched morph (cryptic) is common throughout its range (central and western Europe), while a melanistic morph is frequently found in montane populations, presumably for thermoregulatory reasons. Besides, rare atypical uniformly colored individuals are known here and there. Nevertheless, we found in a restricted treeless area of the French Alps, a population containing a high proportion (>50%) of such specimens. The aim of the study is to bring insight into the presence and function of this color morph by (i) studying the genetic structure of these populations using nine microsatellite markers, and testing for (ii) a potential local diversifying selection and (iii) differences in dispersal capacity between blotched and non-blotched vipers. RESULTS: Our genetic analyses support the occurrence of local diversifying selection for the non-blotched phenotype. In addition, we found significant color-biased dispersal, blotched individuals dispersing more than atypical individuals. CONCLUSION: We hypothesize that, in this population, the non-blotched phenotype possess an advantage over the typical one, a phenomenon possibly due to a better background matching ability in a more open habitat. In addition, color-biased dispersal might be partly associated with the observed local diversifying selection, as it can affect the genetic structure of populations, and hence the distribution of color morphs.

Low-level signaling generated by FcgammaRIIB-B cell receptor co-ligation establishes a state of global B cell receptor nonresponsiveness.

In addition to the stimulatory, antigen-specific B cell receptor (BCR), B lymphocytes also express multiple inhibitory receptors, including Fc gamma receptor type IIB (FcgammaRIIB). Moreover, many laboratories have demonstrated that co-ligation of BCR molecules to inhibitory FcgammaRIIB molecules with high concentrations (10-15 microg/ml) of ligand results in altered BCR signaling. However, there are no reports on the effect of low concentrations of ligand on BCR-FcgammaRIIB co-ligation and subsequent signaling. This knowledge will be critical for optimizing the in vivo use of such reagents. Accordingly, the effect of low ligand concentration on the level of BCR-FcgammaRIIB co-ligation and subsequent BCR signaling was analyzed. The results demonstrate that co-ligation of BCR and FcgammaRIIB molecules at low concentrations (0.5-1.5 microg/ml) of cross-linking reagent, establishes a condition that prevents the B cell from responding to subsequent stimulation, even when the initial exposure to cross-linking reagent fails to generate a calcium flux. Moreover, analysis of the effect of BCR-FcgammaRIIB co-ligation in cells expressing a nonsignaling competent BCR suggest that FcgammaRIIB-mediated inhibition of BCR signaling requires co-ligation of FcgammaRIIB with signaling competent BCR molecules. These results suggest that in vivo treatments with low levels of BCR-FcgammaRIIB cross-linking reagent can induce BCR-FcgammaRIIB co-ligation and establish a condition of B cell nonresponsiveness.