MicroRNAs in B-cells: from normal differentiation to treatment of malignancies.

MicroRNAs (miRNAs) are small non-coding RNAs that play important post-transcriptional regulatory roles in a wide range of biological processes. They are fundamental to the normal development of cells, and evidence suggests that the deregulation of specific miRNAs is involved in malignant transformation due to their function as oncogenes or tumor suppressors. We know that miRNAs are involved in the development of normal B-cells and that different B-cell subsets express specific miRNA profiles according to their degree of differentiation.

Stable phenotype of B-cell subsets following cryopreservation and thawing of normal human lymphocytes stored in a tissue biobank.

Background: Cryopreservation is an acknowledged procedure to store vital cells for future biomarker analyses. Few studies, however, have analyzed the impact of the cryopreservation on phenotyping.

Methods: We have performed a controlled comparison of cryopreserved and fresh cellular aliquots prepared from individual healthy donors.

Stable Phenotype Of B-Cell Subsets Following Cryopreservation and Thawing of Normal Human Lymphocytes Stored in a Tissue Biobank.

Background Cryopreservation is an acknowledged procedure to store vital cells for future biomarker analyses. Few studies, however, have analyzed the impact of the cryopreservation on phenotyping. Methods We have performed a controlled comparison of cryopreserved and fresh cellular aliquots prepared from individual healthy donors.

Human B-cell cancer cell lines as a preclinical model for studies of drug effect in diffuse large B-cell lymphoma and multiple myeloma.

Drug resistance in cancer refers to recurrent or primary refractory disease following drug therapy. At the cellular level, it is a consequence of molecular functions that ultimately enable the cell to resist cell death-one of the classical hallmarks of cancer. Thus, drug resistance is a fundamental aspect of the cancer cell phenotype, in parallel with sustained proliferation, immortality, angiogenesis, invasion, and metastasis.

Exposure time independent summary statistics for assessment of drug dependent cell line growth inhibition.

Background: In vitro generated dose-response curves of human cancer cell lines are widely used to develop new therapeutics. The curves are summarised by simplified statistics that ignore the conventionally used dose-response curves' dependency on drug exposure time and growth kinetics. This may lead to suboptimal exploitation of data and biased conclusions on the potential of the drug in question.

Validation and implementation of a method for microarray gene expression profiling of minor B-cell subpopulations in man.

Background: This report describes a method for the generation of global gene expression profiles from low frequent B-cell subsets by using fluorescence-activated cell sorting and RNA amplification. However, some of the differentiating compartments involve a low number of cells and therefore it is important to optimize and validate each step in the procedure.

Methods: Normal lymphoid tissues from blood, tonsils, thymus and bone marrow were immunophenotyped by the 8-colour Euroflow panel using multiparametric flow cytometry.

Cell of origin associated classification of B-cell malignancies by gene signatures of the normal B-cell hierarchy.

Recent findings have suggested biological classification of B-cell malignancies as exemplified by the "activated B-cell-like" (ABC), the "germinal-center B-cell-like" (GCB) and primary mediastinal B-cell lymphoma (PMBL) subtypes of diffuse large B-cell lymphoma and "recurrent translocation and cyclin D" (TC) classification of multiple myeloma. Biological classification of B-cell derived cancers may be refined by a direct and systematic strategy where identification and characterization of normal B-cell differentiation subsets are used to define the cancer cell of origin phenotype. Here we propose a strategy combining multiparametric flow cytometry, global gene expression profiling and biostatistical modeling to generate B-cell subset specific gene signatures from sorted normal human immature, naive, germinal centrocytes and centroblasts, post-germinal memory B-cells, plasmablasts and plasma cells from available lymphoid tissues including lymph nodes, tonsils, thymus, peripheral blood and bone marrow.

Reproducible probe-level analysis of the Affymetrix Exon 1.0 ST array with R/Bioconductor.

The presence of different transcripts of a gene across samples can be analysed by whole-transcriptome microarrays. Reproducing results from published microarray data represents a challenge owing to the vast amounts of data and the large variety of preprocessing and filtering steps used before the actual analysis is carried out. To guarantee a firm basis for methodological development where results with new methods are compared with previous results, it is crucial to ensure that all analyses are completely reproducible for other researchers.

Multiparametric flow cytometry for identification and fluorescence activated cell sorting of five distinct B-cell subpopulations in normal tonsil tissue.

The purpose of this study was to establish a procedure capable of isolating distinct B-cell subpopulations from human tonsils as a basis for subsequent molecular analyses. Overall, 5 distinct B-cell subpopulations were purified from fresh tonsils based on their fluorescence surface marker expression: naive B cells, centroblasts, centrocytes, memory B cells, and plasmablasts. The immunophenotypic identity of the subpopulations was verified by quantitative real-time reverse transcriptase-polymerase chain reaction using the proliferation marker MKI-67 and 6 B-cell-associated differentiation markers (BACH2, BCL6, PAX5, IRF4, PRDM1, and XBP1).

Cancer stem cells and the cellular hierarchy in haematological malignancies.

Malignancies in the haematopoietic system seem to depend on a small subset of so-called cancer stem cells (CSC) for their continued growth and progression - this was first described as the "sleeper-feeder theory" for leukaemia. The leukaemia stem cell was the first of such subsets to be described although the origins of these cells have been difficult to dissect. Consequently, their biology is not fully elucidated, which also holds true for the normal-tissue counterparts.