CD19 is essential for B cell activation by promoting B cell receptor-antigen microcluster formation in response to membrane-bound ligand.

Here we describe the spatiotemporal architecture, at high molecular resolution, of receptors and signaling molecules during the early events of mouse B cell activation. In response to membrane-bound ligand stimulation, antigen aggregation occurs in B cell antigen receptor (BCR) microclusters containing immunoglobulin (Ig) M and IgD that recruit the kinase Syk and transiently associate with the coreceptor CD19. Unexpectedly, CD19-deficient B cells were significantly defective in initiation of BCR-dependent signaling, accumulation of downstream effectors and cell spreading, defects that culminated in reduced microcluster formation. Hence, we have defined the dynamics of assembly of the main constituents of the BCR 'signalosome' and revealed an essential role for CD19, independent of the costimulatory molecule CD21, in amplifying early B cell activation events in response to membrane-bound ligand stimulation.

The Impact of the Electronic Health Record on Moving New Evidence-Based Nursing Practices Forward.

BACKGROUND: Anecdotal reports from across the country highlight the fact that nurses are facing major challenges in moving new evidence-based practice (EBP) initiatives into the electronic health record (EHR). PURPOSE: The purpose of this study was to: (a) learn current processes for embedding EBP into EHRs, (b) uncover facilitators and barriers associated with rapid movement of new evidence-based nursing practices into the EHR and (c) identify strategies and processes that have been successfully implemented in healthcare organizations across the nation. METHODS: A qualitative study design was utilized. Purposive sampling was used to recruit nurses from across the country (N = 29). Nine focus group sessions were conducted. Semistructured interview questions were developed. Focus groups were conducted by video and audio conferencing. Using an inductive approach, each transcript was read and initial codes were generated resulting in major themes and subthemes. RESULTS: Five major themes were identified: (a) barriers to advancing EBP secondary to the EHR, (b) organizational structure and governing processes of the EHR, (c) current processes for prioritization of EHR changes, (d) impact on ability of clinicians to implement EBP and (e) wait times and delays. LINKING EVIDENCE TO ACTION: Delays in moving new EBP practice changes into the EHR are significant. These delays are sources of frustration and job dissatisfaction. Our results underscore the importance of a priori planning for anticipated changes and building expected delays into the timeline for EBP projects. Moreover, nurse executives must advocate for greater representation of nursing within informatics technology governance structures and additional resources to hire nurse informaticians.

The membrane skeleton controls diffusion dynamics and signaling through the B cell receptor.

Early events of B cell activation after B cell receptor (BCR) triggering have been well characterized. However, little is known about the steady state of the BCR on the cell surface. Here, we simultaneously visualize single BCR particles and components of the membrane skeleton. We show that an ezrin- and actin-defined network influenced steady-state BCR diffusion by creating boundaries that restrict BCR diffusion. We identified the intracellular domain of Igbeta as important in mediating this restriction in diffusion. Importantly, alteration of this network was sufficient to induce robust intracellular signaling and concomitant increase in BCR mobility. Moreover, by using B cells deficient in key signaling molecules, we show that this signaling was most probably initiated by the BCR. Thus, our results suggest the membrane skeleton plays a crucial function in controlling BCR dynamics and thereby signaling, in a way that could be important for understanding tonic signaling necessary for B cell development and survival.

Melting barriers to faunal exchange across ocean basins.

Accelerated loss of sea ice in the Arctic is opening routes connecting the Atlantic and Pacific Oceans for longer periods each year. These changes may increase the ease and frequency with which marine birds and mammals move between the Pacific and Atlantic Ocean basins. Indeed, recent observations of birds and mammals suggest these movements have intensified in recent decades. Reconnection of the Pacific and Atlantic Ocean basins will present both challenges to marine ecosystem conservation and an unprecedented opportunity to examine the ecological and evolutionary consequences of interoceanic faunal exchange in real time. To understand these changes and implement effective conservation of marine ecosystems, we need to further develop modeling efforts to predict the rate of dispersal and consequences of faunal exchange. These predictions can be tested by closely monitoring wildlife dispersal through the Arctic Ocean and using modern methods to explore the ecological and evolutionary consequences of these movements.

Outcomes of an Oral Care Protocol in Postmechanically Ventilated Patients.

BACKGROUND: Oral care is standard practice to prevent hospital-associated infections while patients are intubated and in the intensive care unit. Following extubation and transfer, infections remain an important risk for patients, but less attention is paid to oral care. Few studies have assessed the impact of oral care in recently extubated acutely ill patients. AIMS: To develop an evidence-based oral care protocol for hospitalized patients and determine the impact of this protocol on health outcomes in recently extubated patients. METHODS: In this randomized controlled trial, subjects were randomized to usual care or an intervention protocol that included tooth brushing, tongue scraping, flossing, mouth rinsing, and lip care. Major outcome measures were the revised THROAT (R-THROAT; oral cavity assessment) and overall prevalence of methicillin-sensitive Staphylococcus aureus and methicillin-resistant S. aureus on oral cultures. RESULTS: Seventy-four subjects were randomized. As measured by the R-THROAT, oral cavity health improved over time in both groups, but the intervention group demonstrated significantly more improvement than the control group (R-THROAT score improved by 1.97 intervention vs. 0.87 control; p = .04). Two categories, tongue and mouth comfort, demonstrated the most significant improvement. There was no difference in MSSA/MRSA colonization between the groups at the conclusion of the study. Overall, subjects in the intervention group were more satisfied with their protocol than subjects in the usual care group. LINKING EVIDENCE TO ACTION: This study offers an important evaluation of an oral care protocol after extubation. Results demonstrated improvement in the oral cavity assessment with the designed oral care protocol. Patients expressed a preference for the intervention protocol, which included a battery-operated toothbrush, higher-quality toothpaste and mouth rinse, tongue scraper, floss, and lip balm. The implementation of an oral care protocol specifically addressing patients in the immediate postintubation is essential.

Langerhans cell maturation is accompanied by induction of N-cadherin and the transcriptional regulators of epithelial-mesenchymal transition ZEB1/2.

Langerhans cells (LCs) are a unique subset of dendritic cells (DCs) that express epithelial adhesion molecules, allowing them to form contacts with epithelial cells and reside in epidermal/epithelial tissues. The dynamic regulation of epithelial adhesion plays a decisive role in the life cycle of LCs. It controls whether LCs remain immature and sessile within the epidermis or mature and egress to initiate immune responses. So far, the molecular machinery regulating epithelial adhesion molecules during LC maturation remains elusive. Here, we generated pure populations of immature human LCs in vitro to systematically probe for gene-expression changes during LC maturation. LCs down-regulate a set of epithelial genes including E-cadherin, while they upregulate the mesenchymal marker N-cadherin known to facilitate cell migration. In addition, N-cadherin is constitutively expressed by monocyte-derived DCs known to exhibit characteristics of both inflammatory-type and interstitial/dermal DCs. Moreover, the transcription factors ZEB1 and ZEB2 (ZEB is zinc-finger E-box-binding homeobox) are upregulated in migratory LCs. ZEB1 and ZEB2 have been shown to induce epithelial-to-mesenchymal transition (EMT) and invasive behavior in cancer cells undergoing metastasis. Our results provide the first hint that the molecular EMT machinery might facilitate LC mobilization. Moreover, our study suggests that N-cadherin plays a role during DC migration.

Accuracy of Flash Glucose Monitoring in Hemodialysis Patients With and Without Diabetes Mellitus.

AIMS: Glucose and insulin metabolism are altered in hemodialysis patients, and diabetes management is difficult in these patients. We aimed to validate flash glucose monitoring (FGM) in hemodialysis patients with and without diabetes mellitus as an attractive option for glucose monitoring not requiring regular self-punctures. METHODS: We measured interstitial glucose using a FreeStyle Libre device in eight hemodialysis patients with and seven without diabetes mellitus over 14 days and compared the results to simultaneously performed self-monitoring of capillary blood glucose (SMBG). RESULTS: In 720 paired measurements, mean flash glucose values were significantly lower than self-measured capillary values (6.17±2.52 vs. 7.15±2.41 mmol/L, p=1.3 E-86). Overall, the mean absolute relative difference was 17.4%, and the mean absolute difference was 1.20 mmol/L. The systematic error was significantly larger in patients without vs. with diabetes (- 1.17 vs. - 0.82 mmol/L) and on dialysis vs. interdialytic days (-1.09 vs. -0.90 mmol/L). Compared to venous blood glucose (72 paired measurements), the systematic error of FGM was even larger (5.89±2.44 mmol/L vs. 7.78±7.25 mmol/L, p=3.74E-22). Several strategies to reduce the systematic error were evaluated, including the addition of +1.0 mmol/L as a correction term to all FGM values, which significantly improved accuracy. CONCLUSIONS: FGM systematically underestimates blood glucose in hemodialysis patients but, taking this systematic error into account, the system may be useful for glucose monitoring in hemodialysis patients with or without diabetes.

Serious Illness Discussion in Palliative Care-A Case Study Approach in an African American Patient with Cancer.

Medicare's new focus on end-of-life care has driven nurses and other clinicians to re-examine when advanced care planning should begin, and serious illness discussions should be conducted. This article will address barriers to, cultural influences on, framing of, and documentation of serious illness discussions using a case study approach.

Implementation of a Protocol for Peripheral Intravenous Norepinephrine: Does It Save Central Line Insertion, Is It Safe?

BACKGROUND: Central venous catheters (CVC) are generally recommended for norepinephrine administration due to risk of tissue ischemia. Early resuscitation, leading to decreased infusion duration, may minimize the need for CVCs if norepinephrine can be administered safely through a peripheral intravenous catheter (PIV). OBJECTIVE: A protocol was developed for peripheral administration. Safety, CVC placement, and adherence with protocol elements were evaluated. METHODS: A single-center, prospective, observational pilot was conducted for patients receiving norepinephrine in the Medical Intensive Care Unit (MICU). Patients were considered for PIV administration of low dose norepinephrine for less than 24 hours based on clinical status and anticipated short-term use. Protocolized interventions for PIV's included criteria for gauge, number, and site as well as visual inspection and evaluation every 2 hours. Data was collected on protocol elements to evaluate safety and effectiveness of the protocol. RESULTS: There were 316 occurrences of norepinephrine infusions including 92 via PIV (patients may have received multiple treatments). 34% (31/92) did not require a CVC. 3 had infiltrated PIV's without tissue injury. Maximum dose adherence was 73%. 97% of infusions ran less than 24 hours. Nursing adherence included: 91% gauge, 65% proper site, 99% adequate number, 49% blood return on initiation, 55% ongoing blood return, and 61% IV site checked. CONCLUSION: Our results suggest that norepinephrine is safe to administer through a PIV at low doses for less than 24 hours using a protocol. Prevention of unnecessary CVC insertion is beneficial by minimizing the risk of central line complications thus improving patient morbidity.

The role of the endolithic alga Ostreobium spp. during coral bleaching recovery.

In this study, we explore how the Caribbean coral Orbicella faveolata recovers after bleaching, using fragments from 13 coral colonies exposed to heat stress (32 °C) for ten days. Biological parameters and coral optical properties were monitored during and after the stress. Increases in both, the excitation pressure over photosystem II (Qm) and pigment specific absorption (a*Chla) were observed in the stressed corals, associated with reductions in light absorption at the chlorophyll a red peak (De675) and symbiont population density. All coral fragments exposed to heat stress bleached but a fraction of the stressed corals recovered after removing the stress, as indicated by the reductions in Qm and increases in De675 and the symbiont population observed. This subsample of the experimentally bleached corals also showed blooms of the endolithic algae Ostreobium spp. underneath the tissue. Using a numerical model, we quantified the amount of incident light reflected by the coral, and absorbed by the different pigmented components: symbionts, host-tissue and Ostreobium spp. Our study supports the key contribution of Ostreobium spp. blooms near the skeletal surface, to coral recovery after bleaching by reducing skeleton reflectance. Endolithic blooms can thus significantly alleviate the high light stress that affects the remaining symbionts during the stress or when the coral has achieved the bleached phenotype.

Complex Oncologic Surgeries and Implications for the Intensive Care Unit Nurse.

There are many challenges in caring for the postsurgical patient in the intensive care unit. When the postsurgical patient has an active malignancy, this can make the intensive care unit care more challenging. Nutrition, infection, and the need for postoperative mechanical ventilatory support for the patient with cancer present challenges that may increase the patient's length of stay in the intensive care unit. Critical care nurses must be aware of these challenges as they provide care to this patient population.

Approach for tuberculosis-associated immune reconstitution inflammatory syndrome in an HIV-negative patient.

A male refugee from the Middle East was diagnosed with pulmonary tuberculosis and Pott's disease with paravertebral abscess. After starting the standard regimen, the sputum culture converted to negative and the patient's general condition improved. Six weeks later, the patient presented with clinical worsening of known symptoms, new appearance of focal neurological deficits and progress of radiological features showing progression of the paravertebral abscess. Immune reconstitution inflammatory syndrome with Mycobacterium tuberculosis (TB-IRIS) was presumed, and treatment with high-dose steroids was started. Due to recurrent relapses while tapering, corticosteroids had to be given over a prolonged period. After treatment completion, the patient was in a good general condition, abscesses had decreased and neurological deficits were in complete remission. This case presents the rare manifestation of TB-IRIS in HIV-negative patients and its management in a high-income country.

Wiskott-Aldrich syndrome protein deficiency in B cells results in impaired peripheral homeostasis.

To more precisely identify the B-cell phenotype in Wiskott-Aldrich syndrome (WAS), we used 3 distinct murine in vivo models to define the cell intrinsic requirements for WAS protein (WASp) in central versus peripheral B-cell development. Whereas WASp is dispensable for early bone marrow B-cell development, WASp deficiency results in a marked reduction in each of the major mature peripheral B-cell subsets, exerting the greatest impact on marginal zone and B1a B cells. Using in vivo bromodeoxyuridine labeling and in vitro functional assays, we show that these deficits reflect altered peripheral homeostasis, partially resulting from an impairment in integrin function, rather than a developmental defect. Consistent with these observations, we also show that: (1) WASp expression levels increase with cell maturity, peaking in those subsets exhibiting the greatest sensitivity to WASp deficiency; (2) WASp(+) murine B cells exhibit a marked selective advantage beginning at the late transitional B-cell stage; and (3) a similar in vivo selective advantage is manifest by mature WASp(+) human B cells. Together, our data provide a better understanding of the clinical phenotype of WAS and suggest that gene therapy might be a useful approach to rescue altered B-cell homeostasis in this disease.

The chemokine receptor D6 constitutively traffics to and from the cell surface to internalize and degrade chemokines.

The D6 heptahelical membrane protein, expressed by lymphatic endothelial cells, is able to bind with high affinity to multiple proinflammatory CC chemokines. However, this binding does not allow D6 to couple to the signaling pathways activated by typical chemokine receptors such as CC-chemokine receptor-5 (CCR5). Here, we show that D6, like CCR5, can rapidly internalize chemokines. However, D6-internalized chemokines are more effectively retained intracellularly because they more readily dissociate from the receptor during vesicle acidification. These chemokines are then degraded while the receptor recycles to the cell surface. Interestingly, D6-mediated chemokine internalization occurs without bringing about a reduction in cell surface D6 levels. This is possible because unlike CCR5, D6 is predominantly localized in recycling endosomes capable of trafficking to and from the cell surface in the absence of ligand. When chemokine is present, it can enter the cells associated with D6 already destined for internalization. By this mechanism, D6 can target chemokines for degradation without the necessity for cell signaling, and without desensitizing the cell to subsequent chemokine exposure.

Knockdown of mouse VCAM-1 by vector-based siRNA.

Graft rejection is critically dependent on the recruitment of leukocytes via adhesion molecules on the endothelium, and inhibition of these interactions can prolong graft survival. We have therefore developed an approach using siRNA to inhibit the expression of VCAM-1 in endothelial cells. We transfected siRNA constructs into murine corneal and vascular endothelium and looked at expression of VCAM-1 and other surface molecules by flow cytometry. Adhesion assays (both static and under flow) were used to determine the effect of VCAM-1 inhibition. The activation of cellular stress responses was assessed by RT-PCR. Constructs encoding siRNA can block expression of VCAM-1 in both corneal and vascular endothelial cells (in the latter case after cytokine stimulation). Inhibition of VCAM-1 expression reduced the ability of T cells to adhere to endothelium. However, there were non-specific effects of siRNA expression, including upregulation of (Programmed Death Ligand 1) PDL1 and decreased cell growth. Analysis of stress pathways showed that the endothelial cells transfected with siRNA had upregulated molecules associated with cell stress. While these data are supportive of a potential therapeutic role for siRNA constructs in blocking the expression of adhesion molecules, they also highlight potential non-specific effects of siRNA that must be carefully considered in any application of this technology.

Antimicrobial resistance data on 16,756 Streptococcus pneumoniae isolates in 1999: A Pan-Regional Multicenter Surveillance Study in France.

The rising prevalence of antibiotic-resistant Streptococcus pneumoniae is a phenomenon observed to different degrees around the world. The present national surveillance study report analyzes a total of 16,756 strains of S. pneumoniae collected across France in 1999. The overall prevalence of S. pneumoniae with decreased susceptibility to penicillin was 44%, to amoxicillin 26%, and to cefotaxime 17%. The proportion of high-level resistant strains to penicillin (MIC > 1 mg/L), amoxicillin and cefotaxime (MIC > 2 mg/L) remained low: 12.3%, 1.8%, and 0.4% respectively. Prevalence of resistance to other antibiotics was high: 53% to erythromycin, 41.7% to cotrimoxazole, 31.8% to tetracycline, and 24.6% to chloramphenicol. Prevalence of penicillin-resistant S. pneumoniae varied according to subject age and specimen source. It was higher in children (52.7%) than in adults (39.8%) and higher in strains isolated from middle ear fluid (63.6%) than from blood cultures (41.8%) in children. S. pneumoniae resistant to other antibiotics were more common in children than in adults, although figures showed geographical variations. Comparison with a previous study realized in 1997 in the same regions confirms a rising trend in the prevalence of resistant bacteria. Therefore, we conclude that prevalence of antibiotic-resistant S. pneumoniae in 1999 continued to rise in France, although strains with high-level resistance to penicillin remained stable.

Endocarditis in patients with a permanent pacemaker: a 1-year epidemiological survey on infective endocarditis due to valvular and/or pacemaker infection.

To describe characteristics of infective endocarditis (IE) in pacemaker (PM) recipients, including the annual incidence and exact localization of IE on PM leads, cardiac valves, or both, we prospectively analyzed 45 PM recipients from a group of 559 patients with definite IE who responded to a population-based survey conducted in France in 1999. Thirty-three patients had definite PM-lead IE (group I), and 12 had valvular IE without evidence of PM involvement (group II). The valvular structure was involved in almost two-thirds of IE cases among PM recipients. Of the 28 patients (62%) with valvular IE, 10 group I patients had tricuspid involvement, and 6 group I patients had left heart-valve involvement. The most frequent causative organisms in groups I and II were staphylococci (82%) and streptococci (50%), respectively. The incidence of age- and sex-standardized IE was 550 cases/million PM recipients per year. The incidence of IE with PM involvement is between that of valvular IE in the general population and prosthetic valve IE.

Coral transcriptome and bacterial community profiles reveal distinct Yellow Band Disease states in Orbicella faveolata.

Coral diseases impact reefs globally. Although we continue to describe diseases, little is known about the etiology or progression of even the most common cases. To examine a spectrum of coral health and determine factors of disease progression we examined Orbicella faveolata exhibiting signs of Yellow Band Disease (YBD), a widespread condition in the Caribbean. We used a novel combined approach to assess three members of the coral holobiont: the coral-host, associated Symbiodinium algae, and bacteria. We profiled three conditions: (1) healthy-appearing colonies (HH), (2) healthy-appearing tissue on diseased colonies (HD), and (3) diseased lesion (DD). Restriction fragment length polymorphism analysis revealed health state-specific diversity in Symbiodinium clade associations. 16S ribosomal RNA gene microarrays (PhyloChips) and O. faveolata complimentary DNA microarrays revealed the bacterial community structure and host transcriptional response, respectively. A distinct bacterial community structure marked each health state. Diseased samples were associated with two to three times more bacterial diversity. HD samples had the highest bacterial richness, which included components associated with HH and DD, as well as additional unique families. The host transcriptome under YBD revealed a reduced cellular expression of defense- and metabolism-related processes, while the neighboring HD condition exhibited an intermediate expression profile. Although HD tissue appeared visibly healthy, the microbial communities and gene expression profiles were distinct. HD should be regarded as an additional (intermediate) state of disease, which is important for understanding the progression of YBD.

Live cell imaging of chemotactic dendritic cell migration in explanted mouse ear preparations.

Leukocyte migration through the interstitial space is crucial for the maintenance of tolerance and immunity. The main cues for leukocyte trafficking are chemokines thought to directionally guide these cells towards their targets. However, model systems that facilitate quantification of chemokine-guided leukocyte migration in vivo are uncommon. Here we describe an ex vivo crawl-in assay using explanted mouse ears that allows the visualization of chemokine-dependent dendritic cell (DC) motility in the dermal interstitium in real time. We present methods for the preparation of mouse ear sheets and their use in multidimensional confocal imaging experiments to monitor and analyze the directional migration of fluorescently labelled DCs through the dermis and into afferent lymphatic vessels. The assay provides a more physiological approach to study leukocyte migration than in vitro three-dimensional (3D) or 2-dimensional (2D) migration assays such as collagen gels and transwell assays.