Affinity flow fractionation of cells via transient interactions with asymmetric molecular patterns

Flow fractionation of cells using physical fields to achieve lateral displacement finds wide applications, but its extension to surface molecule-specific separation requires labeling. Here we demonstrate affinity flow fractionation (AFF) where weak, short-range interactions with asymmetric molecular patterns laterally displace cells in a continuous, label-free process. We show that AFF can directly draw neutrophils out of a continuously flowing stream of blood with an unprecedented 400,000-fold depletion of red blood cells, with the sorted cells being highly viable, unactivated, and functionally intact. The lack of background erythrocytes enabled the use of AFF for direct enumeration of neutrophils by a downstream detector, which could distinguish the activation state of neutrophils in blood. The compatibility of AFF with capillary microfluidics and its ability to directly separate cells with high purity and minimal sample preparation will facilitate the design of simple and portable devices for point-of-care diagnostics and quick, cost-effective laboratory analysis

A Large Web-Based Observer Reliability Study of Early Ischaemic Signs on Computed Tomography. The Acute Cerebral CT Evaluation of Stroke Study (ACCESS)

BACKGROUND: Early signs of ischaemic stroke on computerised tomography (CT) scanning are subtle but CT is the most widely available diagnostic test for stroke. Scoring methods that code for the extent of brain ischaemia may improve stroke diagnosis and quantification of the impact of ischaemia. METHODOLOGY AND PRINCIPAL FINDINGS: We showed CT scans from patients with acute ischaemic stroke (n = 32, with different patient characteristics and ischaemia signs) to doctors in stroke-related specialties world-wide over the web. CT scans were shown twice, randomly and blindly. Observers entered their scan readings, including early ischaemic signs by three scoring methods, into the web database. We compared observers' scorings to a reference standard neuroradiologist using area under receiver operator characteristic curve (AUC) analysis, Cronbach's alpha and logistic regression to determine the effect of scales, patient, scan and observer variables on detection of early ischaemic changes. Amongst 258 readers representing 33 nationalities and six specialties, the AUCs comparing readers with the reference standard detection of ischaemic signs were similar for all scales and both occasions. Being a neuroradiologist, slower scan reading, more pronounced ischaemic signs and later time to CT all improved detection of early ischaemic signs and agreement on the rating scales. Scan quality, stroke severity and number of years of training did not affect agreement. CONCLUSIONS: Large-scale observer reliability studies are possible using web-based tools and inform routine practice. Slower scan reading and use of CT infarct rating scales improve detection of acute ischaemic signs and should be encouraged to improve stroke diagnosis

Fluctuation theorem for currents and Schnakenberg network theory

A fluctuation theorem is proved for the macroscopic currents of a system in a nonequilibrium steady state, by using Schnakenberg network theory. The theorem can be applied, in particular, in reaction systems where the affinities or thermodynamic forces are defined globally in terms of the cycles of the graph associated with the stochastic process describing the time evolution.Comment: new version : 16 pages, 1 figure, to be published in Journal of Statistical Physic

TIM-1 and TIM-4 Glycoproteins Bind Phosphatidylserine and Mediate Uptake of Apoptotic Cells

SummaryThe T cell immunoglobulin mucin (TIM) proteins regulate T cell activation and tolerance. Here we showed that TIM-4 is expressed on human and mouse macrophages and dendritic cells, and both TIM-4 and TIM-1 specifically bound phosphatidylserine (PS) on the surface of apoptotic cells but not any other phospholipid tested. TIM-4+ peritoneal macrophages, TIM-1+ kidney cells, and TIM-4- or TIM-1-transfected cells efficiently phagocytosed apoptotic cells, and phagocytosis could be blocked by TIM-4 or TIM-1 monoclonal antibodies. Mutations in the unique cavity of TIM-4 eliminated PS binding and phagocytosis. TIM-4 mAbs that blocked PS binding and phagocytosis mapped to epitopes in this binding cavity. These results show that TIM-4 and TIM-1 are immunologically restricted members of the group of receptors whose recognition of PS is critical for the efficient clearance of apoptotic cells and prevention of autoimmunity

Leadership Styles and Innovative Entrepreneurship: An International Study

This research attempts to empirically examine the relationship between leadership styles and innovative entrepreneurship through regression analysis, using a sample of 43 countries and data from Global Entrepreneurship Monitor and Global Leadership and Organizational Behavior Effectiveness. In light of institutional approaches and specifically based on the normative dimension, the main findings of the study indicate that participative leadership and higher education represent the strongest explanatory factor in the variance of the current rates of innovative entrepreneurship. This study has contributions for both researchers and policymakers on new firm creation (entrepreneurship) and on the generation of innovation within organizations (intrapreneurship).Stefan van Hemmen acknowledges the financial resources from ECO2013-48496-C4-4-R (Spanish Ministry of Economy & Competitiveness) and 2014-SGR-1259 (Economy & Knowledge Department -Catalan Government-). Marta Peris-Ortiz acknowledges support from the Universitat Politecnica de Valencia through the project Paid-06-12 (Sp 20120792). Claudia Alvarez and David Urbano acknowledge the financial support from the Projects ECO2013-44027-P (Spanish Ministry of Economy & Competitiveness) and 2014-SGR-1626)Economy & Knowledge Department -Catalan Government-).Van Hemmen, S.; Alvarez, C.; Peris-Ortiz, M.; Urbano, D. (2015). Leadership Styles and Innovative Entrepreneurship: An International Study. Cybernetics and Systems. 46(3-4):271-286. https://doi.org/10.1080/01969722.2015.1012896S271286463-

Gauging NOTCH1 Activation in Cancer Using Immunohistochemistry

Fixed, paraffin-embedded (FPE) tissues are a potentially rich resource for studying the role of NOTCH1 in cancer and other pathologies, but tests that reliably detect activated NOTCH1 (NICD1) in FPE samples have been lacking. Here, we bridge this gap by developing an immunohistochemical (IHC) stain that detects a neoepitope created by the proteolytic cleavage event that activates NOTCH1. Following validation using xenografted cancers and normal tissues with known patterns of NOTCH1 activation, we applied this test to tumors linked to dysregulated Notch signaling by mutational studies. As expected, frequent NICD1 staining was observed in T lymphoblastic leukemia/lymphoma, a tumor in which activating NOTCH1 mutations are common. However, when IHC was used to gauge NOTCH1 activation in other human cancers, several unexpected findings emerged. Among B cell tumors, NICD1 staining was much more frequent in chronic lymphocytic leukemia than would be predicted based on the frequency of NOTCH1 mutations, while mantle cell lymphoma and diffuse large B cell lymphoma showed no evidence of NOTCH1 activation. NICD1 was also detected in 38% of peripheral T cell lymphomas. Of interest, NICD1 staining in chronic lymphocytic leukemia cells and in angioimmunoblastic lymphoma was consistently more pronounced in lymph nodes than in surrounding soft tissues, implicating factors in the nodal microenvironment in NOTCH1 activation in these diseases. Among carcinomas, diffuse strong NICD1 staining was observed in 3.8% of cases of triple negative breast cancer (3 of 78 tumors), but was absent from 151 non-small cell lung carcinomas and 147 ovarian carcinomas. Frequent staining of normal endothelium was also observed; in line with this observation, strong NICD1 staining was also seen in 77% of angiosarcomas. These findings complement insights from genomic sequencing studies and suggest that IHC staining is a valuable experimental tool that may be useful in selection of patients for clinical trials

Expression of Natural Killer Receptor Alleles at Different Ly49 Loci Occurs Independently and Is Regulated by Major Histocompatibility Complex Class I Molecules

Ly49 receptor genes are expressed by subsets of natural killer (NK) cells in an overlapping fashion, accounting for the capacity of NK subsets to attack host cells that have selectively downregulated self–major histocompatibility complex (MHC) class I molecules. It was shown previously that most NK cells express only one or the other allele of a given Ly49 gene, while a smaller population expresses both alleles. However, the methods used to detect monoallelic and biallelic cells were nonquantitative. Here, new allele-specific antibodies were used to provide the first quantitative examination of biallelic and monoallelic expression of Ly49A and Ly49G2. The results demonstrate conclusively that most Ly49A+ and Ly49G2+ NK cells express the corresponding gene in a monoallelic fashion, with a smaller subset expressing both alleles. Unexpectedly, biallelic Ly49A+ NK cells were more numerous than predicted by completely independent allelic expression, suggesting some heterogeneity among NK progenitors in the potential to express a given Ly49 gene. The data also show that cells expressing one allele of Ly49G2 may express Ly49A from the same or opposite chromosome with equal likelihood, indicating that the expressed allele is chosen independently for different Ly49 genes. Finally, the data demonstrate that biallelic expression of Ly49A or Ly49G2 occurs least frequently in mice that express ligands for these receptors (H-2d mice), and most frequently in class I–deficient mice. Thus, biallelic expression of Ly49 genes is regulated by interactions of NK cell progenitors with MHC class I molecules

Increase in perceived case suspiciousness due to local contrast optimisation in digital screening mammography

Item does not contain fulltextOBJECTIVES: To determine the influence of local contrast optimisation on diagnostic accuracy and perceived suspiciousness of digital screening mammograms. METHODS: Data were collected from a screening region in the Netherlands and consisted of 263 digital screening cases (153 recalled,110 normal). Each case was available twice, once processed with a tissue equalisation (TE) algorithm and once with local contrast optimisation (PV). All cases had digitised previous mammograms. For both algorithms, the probability of malignancy of each finding was scored independently by six screening radiologists. Perceived case suspiciousness was defined as the highest probability of malignancy of all findings of a radiologist within a case. Differences in diagnostic accuracy of the processing algorithms were analysed by comparing the areas under the receiver operating characteristic curves (A(z)). Differences in perceived case suspiciousness were analysed using sign tests. RESULTS: There was no significant difference in A(z) (TE: 0.909, PV 0.917, P = 0.46). For all radiologists, perceived case suspiciousness using PV was higher than using TE more often than vice versa (ratio: 1.14-2.12). This was significant (P <0.0083) for four radiologists. CONCLUSIONS: Optimisation of local contrast by image processing may increase perceived case suspiciousness, while diagnostic accuracy may remain similar. KEY POINTS: Variations among different image processing algorithms for digital screening mammography are large. Current algorithms still aim for optimal local contrast with a low dynamic range. Although optimisation of contrast may increase sensitivity, diagnostic accuracy is probably unchanged. Increased local contrast may render both normal and abnormal structures more conspicuous.1 april 201

A novel 3D mesenchymal stem cell model of the multiple myeloma bone marrow niche : biologic and clinical applications

Specific niches within the tumor bone marrow (BM) microenvironment afford a sanctuary for multiple myeloma (MM) clones due to stromal cell-tumor cell interactions, which confer survival advantage and drug resistance. Defining the sequelae of tumor cell interactions within the MM niches on an individualized basis may provide the rationale for personalized therapies. To mimic the MM niche, we here describe a new 3D co-culture ex-vivo model in which primary MM patient BM cells are co-cultured with mesenchymal stem cells (MSC) in a hydrogel 3D system. In the 3D model, MSC with conserved phenotype (CD73+CD90+CD105+) formed compact clusters with active fibrous connections, and retained lineage differentiation capacity. Extracellular matrix molecules, integrins, and niche related molecules including N-cadherin and CXCL12 are expressed in 3D MSC model. Furthermore, activation of osteogenesis (MMP13, SPP1, ADAMTS4, and MGP genes) and osteoblastogenic differentiation was confirmed in 3D MSC model. Co-culture of patient-derived BM mononuclear cells with either autologous or allogeneic MSC in 3D model increased proliferation of MM cells, CXCR4 expression, and SP cells. We carried out immune profiling to show that distribution of immune cell subsets was similar in 3D and 2D MSC model systems. Importantly, resistance to novel agents (IMiDs, bortezomib, carfilzomib) and conventional agents (doxorubicin, dexamethasone, melphalan) was observed in 3D MSC system, reflective of clinical resistance. This 3D MSC model may therefore allow for studies of MM pathogenesis and drug resistance within the BM niche. Importantly, ongoing prospective trials are evaluating its utility to inform personalized targeted and immune therapy in MM