Separation of suspended particles in microfluidic systems by directional-locking in periodic fields

We investigate the transport and separation of overdamped particles under the action of a uniform external force in a two-dimensional periodic energy landscape. Exact results are obtained for the deterministic transport in a square lattice of parabolic, repulsive centers that correspond to a piecewise-continuous linear-force model. The trajectories are periodic and commensurate with the obstacle lattice and exhibit phase-locking behavior in that the particle moves at the same average migration angle for a range of orientation of the external force. The migration angle as a function of the orientation of the external force has a Devil's staircase structure. The first transition in the migration angle was analyzed in terms of a Poincare map, showing that it corresponds to a tangent bifurcation. Numerical results show that the limiting behavior for impenetrable obstacles is equivalent to the high Peclet number limit in the case of transport of particles in a periodic pattern of solid obstacles. Finally, we show how separation occurs in these systems depending on the properties of the particles

Commentary on the WHO classification of tumors of lymphoid tissues (2008): “Gray zone” lymphomas overlapping with Burkitt lymphoma or classical Hodgkin lymphoma

The 2008 WHO Classification of Tumors of Haematopoietic and Lymphoid Tissues has introduced two new categories of high-grade B-cell lymphomas: entities in which features of diffuse large B-cell lymphoma (DLBCL) overlap with Burkitt lymphoma (DLBCL/BL) or classical Hodgkin lymphoma (DLBCL/HL). The DLBCL/BL category encompasses cases that resemble Burkitt lymphoma morphologically, but have one or more immunophenotypic or molecular genetic deviations that would exclude it from the BL category; conversely, some cases have immunophenotypic and/or genetic features of BL, but display cytologic variability unacceptable for BL. Many of the cases in the DLBCL/BL category contain a translocation of MYC as well as either BCL2 or BCL6 (so-called double-hit lymphomas) and have a very aggressive clinical behavior. The DLBCL/HL category encompasses lymphomas that exhibit the morphology of classical Hodgkin lymphoma but the immunophenotype of DLBCL, or vice versa. Most DLBCL/HL cases described present as mediastinal masses, but this category is not limited to mediastinal lymphomas. These new categories acknowledge the increasing recognition of cases that display mixed features of two well-established diseases. Whether the existence of such cases reflects shortcomings of our current diagnostic armamentarium or a true disease continuum in which such hybrid or intermediate neoplasms actually exist remains to be determined

Commentary on the WHO classification of tumors of lymphoid tissues (2008): indolent B cell lymphomas

The 4th edition of the World Health Organization classification of tumors of hematopoietic and lymphoid tissues introduces many new items to the classification scheme of the so-called indolent B cell lymphomas. New proposed entities, such as splenic B cell lymphoma/leukemia, unclassifiable, splenic diffuse red pulp small B cell lymphoma, hairy cell leukemia variant, pediatric follicular lymphoma, and pediatric marginal zone lymphoma have been coined, and some definitions of established diseases, such as chronic lymphocytic leukemia or Waldenström’s macroglobulinemia have been revised. One aspect of major importance is the recent description of small clonal B cell populations, in part with a CLL phenotype, and their relationship to B-CLL. Some new subtypes or variants of follicular lymphoma with distinct clinicopathologic and/or molecular genetic characteristics have been described, including primary follicular lymphomas of the duodenum and pediatric follicular lymphomas. Furthermore, the impact of some probably early, or precursor lesions, such as follicular lymphoma in situ is discussed. Overall, we succinctly discuss the essential elements of the revisions made in the updated classification, and we identify potential opportunities for refinement of new or provisional categories in subsequent classifications

The many faces of small B cell lymphomas with plasmacytic differentiation and the contribution of MYD88 testing

Plasmacytic differentiation may occur in almost all small B cell lymphomas (SBLs), although it varies from being uniformly present (as in lymphoplasmacytic lymphoma (LPL)) to very uncommon (as in mantle cell lymphomas (MCLs)). The discovery of MYD88 L265P mutations in the vast majority of LPLs has had a major impact on the study of these lymphomas. Review of the cases contributed to the 2014 European Association for Haematopathology/Society for Hematopathology slide workshop illustrated how mutational testing has helped refine the diagnostic criteria for LPL, emphasizing the importance of identifying a clonal monotonous lymphoplasmacytic population and highlighting how LPL can still be diagnosed with extensive nodal architectural effacement, very subtle plasmacytic differentiation, follicular colonization, or uncommon phenotypes such as CD5 or CD10 expression. MYD88 L265P mutations were found in 11/11 LPL cases versus only 2 of 28 other SBLs included in its differential diagnosis. Mutational testing also helped to exclude other cases that would have been considered LPL in the past. The workshop also highlighted how plasmacytic differentiation can occur in chronic lymphocytic leukemia/small lymphocytic lymphoma, follicular lymphoma, SOX11 negative MCL, and particularly in marginal zone lymphomas, all of which can cause diagnostic confusion with LPL. The cases also highlighted the difficulty in distinguishing lymphomas with marked plasmacytic differentiation from plasma cell neoplasms. Some SBLs with plasmacytic differentiation can be associated with amyloid, other immunoglobulin deposition, or crystal-storing histiocytosis, which may obscure the underlying neoplasm. Finally, although generally indolent, LPL may transform, with the workshop cases suggesting a role for TP53 abnormalities

WNT4 overexpression and secretion in thymic epithelial tumors drive an autocrine loop in tumor cells in vitro

BackgroundWNT4-driven non-canonical signaling is crucial for homeostasis and age-related involution of the thymus. Abnormal WNT signaling is important in many cancers, but the role of WNT signaling in thymic tumors is largely unknown.Materials & MethodsExpression and function of WNT4 and FZD6 were analyzed using qRT–PCR, Western blot, ELISA, in biopsies of non-neoplastic thymi (NT), thymoma and thymic carcinomas. ShRNA techniques and functional assays were used in primary thymic epithelial cells (pTECs) and TC cell line 1889c. Cells were conventionally (2D) grown and in three-dimensional (3D) spheroids.ResultsIn biopsy, WHO classified B3 thymomas and TCs showed increased WNT4 expression compared with NTs. During short-term 2D culture, WNT4 expression and secretion declined in neoplastic pTECs but not in 3D spheroids or medium supplemented with recombinant WNT4 cultures. Under the latter condition, the growth of pTECs was accompanied by increased expression of non-canonical targets RAC1 and JNK. Down-regulation of WNT4 by shRNA induced cell death in pTECs derived from B3 thymomas and led to decreased RAC1, but not JNK protein phosphorylation. Pharmacological inhibition of NF-κB decreased both RAC1 and JNK phosphorylation in neoplastic pTECs.ConclusionsLack of the age-related decline of non-canonical WNT4 expression in TETs and restoration of declining WNT4 expression through exogeneous WNT4 or 3D culture of pTECs hints at an oncogenic role of WNT4 in TETs and is compatible with the WNT4 autocrine loop model. Crosstalk between WNT4 and NF-κB signaling may present a promising target for combined interventions in TETs

Frequent polymorphic changes but not mutations of TRAIL receptors DR4 and DR5 in mantle cell lymphoma and other B-cell lymphoid neoplasms

Background and objectives: tumor necrosis factor related apoptosis-inducing ligand (TRAIL) receptors DR4 and DR5 have been mapped to chromosome 8p21-22, a region frequently deleted in different lymphoid neoplasms. Design and methods: to investigate the potential alterations of these genes in lymphoid neoplasms, we examined the presence of gene mutations in exons 3, 4, and 9 in 69 cases with mantle cell lymphoma (MCL), 16 with chronic lymphocytic leukemia (CLL), 12 with follicular lymphomas (FL) and 17 with large B-cell-lymphomas (DLBCL), as well as in 4 lymphoid cell lines carrying the t(11;14) translocation, and 91 healthy blood donors. Results: three CLL and three MCL cases had 8p deletions. Two nucleotide changes in or near the intron 3 splice consensus sequence and a silent change were found. These rare changes were also present in the germ-line of the patients. The DR4 death domain A1322G polymorphism was significantly more frequent in MCL [odds ratio (OR) = 5.9; 95% confidence interval (CI), 1.92-18.1] and CLL (OR = 4.5; CI, 1.18-17) patients than in a sex and age-adjusted healthy population. In contrast, the DR4 exon 4 C626G polymorphism was associated with a significant overall decreased risk for MCL (OR = 0.3; CI, 0.12-0.8). No mutations or cancer-associated polymorphic changes were found in DR5 domains. Interpretation and conclusions: these findings indicate that mutations of DR4 and DR5 are uncommon in lymphoid neoplasms but DR4 polymorphic alleles may contribute to the pathogenesis of these malignancies

Serum glucose levels for predicting death in patients admitted to hospital for community acquired pneumonia: prospective cohort study

Objective To examine whether acute dysglycaemia predicts death in people admitted to hospital with community acquired pneumonia

Targeted panel sequencing in the routine diagnosis of mature T- and NK-cell lymphomas: report of 128 cases from two German reference centers

Diagnosing any of the more than 30 types of T-cell lymphomas is considered a challenging task for many pathologists and currently requires morphological expertise as well as the integration of clinical data, immunophenotype, flow cytometry and clonality analyses. Even considering all available information, some margin of doubt might remain using the current diagnostic procedures. In recent times, the genetic landscape of most T-cell lymphomas has been elucidated, showing a number of diagnostically relevant mutations. In addition, recent data indicate that some of these genetic alterations might bear prognostic and predictive value. Extensive genetic analyses, such as whole exome or large panel sequencing are still expensive and time consuming, therefore limiting their application in routine diagnostic. We therefore devoted our effort to develop a lean approach for genetic analysis of T-cell lymphomas, focusing on maximum efficiency rather than exhaustively covering all possible targets. Here we report the results generated with our small amplicon-based panel that could be used routinely on paraffin-embedded and even decalcified samples, on a single sample basis in parallel with other NGS-panels used in our routine diagnostic lab, in a relatively short time and with limited costs. We tested 128 available samples from two German reference centers as part of our routine work up (among which 116 T-cell lymphomas), which is the largest routine diagnostic series reported to date. Our results showed that this assay had a very high rate of technical success (97%) and could detect mutations in the majority (79%) of tested T-cell lymphoma samples

Molecular Cytogenetic Profiling Reveals Similarities and Differences Between Localized Nodal and Systemic Follicular Lymphomas

Recently, we have developed novel highly promising gene expression (GE) classifiers discriminating localized nodal (LFL) from systemic follicular lymphoma (SFL) with prognostic impact. However, few data are available in LFL especially concerning hotspot genetic alterations that are associated with the pathogenesis and prognosis of SFL. A total of 144 LFL and 527 SFL, enrolled in prospective clinical trials of the German Low Grade Lymphoma Study Group, were analyzed by fluorescence in situ hybridization to detect deletions in chromosomes 1p, 6q, and 17p as well as BCL2 translocations to determine their impact on clinical outcome of LFL patients. The frequency of chromosomal deletions in 1p and 17p was comparable between LFL and SFL, while 6q deletions and BCL2 translocations more frequently occurred in SFL. A higher proportion of 1p deletions was seen in BCL2-translocation–positive LFL, compared with BCL2-translocation–negative LFL. Deletions in chromosomes 1p, 6q, and 17p predicted clinical outcome of patients with SFL in the entire cohort, while only deletions in chromosome 1p retained its negative prognostic impact in R-CHOP–treated SFL. In contrast, no deletions in one of the investigated genetic loci predicted clinical outcome in LFL. Likewise, the presence or absence of BCL2 translocations had no prognostic impact in LFL. Despite representing a genetic portfolio closely resembling SFL, LFL showed some differences in deletion frequencies. BCL2 translocation and 6q deletion frequency differs between LFL and SFL and might contribute to distinct genetic profiles in LFL and SFL

Cyclin D1-CDK4 activity drives sensitivity to bortezomib in mantle cell lymphoma by blocking autophagy-mediated proteolysis of NOXA

Background: Mantle cell lymphoma (MCL) is an aggressive B-non-Hodgkin lymphoma with generally poor outcome. MCL is characterized by an aberrantly high cyclin D1-driven CDK4 activity. New molecular targeted therapies such as inhibitors of the ubiquitin-proteasome system (UPS) have shown promising results in preclinical studies and MCL patients. Our previous research revealed stabilization of the short-lived pro-apoptotic NOXA as a critical determinant for sensitivity to these inhibitors. It is currently unclear how cyclin D1 overexpression and aberrant CDK4 activity affect NOXA stabilization and treatment efficacy of UPS inhibitors in MCL. Methods: The effect of cyclin D1-driven CDK4 activity on response of MCL cell lines and primary cells to proteasome inhibitor treatment was investigated using survival assays (Flow cytometry, AnnexinV/PI) and Western blot analysis of NOXA protein. Half-life of NOXA protein was determined by cycloheximide treatment and subsequent Western blot analysis. The role of autophagy was analyzed by LC3-II protein expression and autophagolysosome detection. Furthermore, silencing of autophagy-related genes was performed using siRNA and MCL cells were treated with autophagy inhibitors in combination with proteasome and CDK4 inhibition. Results: In this study, we show that proteasome inhibitor-mediated cell death in MCL depends on cyclin D1-driven CDK4 activity. Inhibition of cyclin D1/CDK4 activity significantly reduced proteasome inhibitor-mediated stabilization of NOXA protein, mainly driven by an autophagy-mediated proteolysis. Bortezomib-induced cell death was significantly potentiated by compounds that interfere with autophagosomal function. Combined treatment with bortezomib and autophagy inhibitors enhanced NOXA stability leading to super-induction of NOXA protein. In addition to established autophagy modulators, we identified the fatty acid synthase inhibitor orlistat to be an efficient autophagy inhibitor when used in combination with bortezomib. Accordingly, this combination synergistically induced apoptosis both in MCL cell lines and in patient samples. Conclusion: Our data demonstrate that CDK4 activity in MCL is critical for NOXA stabilization upon treatment with UPS inhibitors allowing preferential induction of cell death in cyclin D transformed cells. Under UPS blocked conditions, autophagy appears as the critical regulator of NOXA induction. Therefore, inhibitors of autophagy are promising candidates to increase the activity of proteasome inhibitors in MCL