A theorem proving approach for automatically synthesizing visualizations of flow cytometry data.

BACKGROUND: Polychromatic flow cytometry is a popular technique that has wide usage in the medical sciences, especially for studying phenotypic properties of cells. The high-dimensionality of data generated by flow cytometry usually makes it difficult to visualize. The naive solution of simply plotting two-dimensional graphs for every combination of observables becomes impractical as the number of dimensions increases. A natural solution is to project the data from the original high dimensional space to a lower dimensional space while approximately preserving the overall relationship between the data points. The expert can then easily visualize and analyze this low-dimensional embedding of the original dataset. RESULTS: This paper describes a new method, SANJAY, for visualizing high-dimensional flow cytometry datasets. This technique uses a decision procedure to automatically synthesize two-dimensional and three-dimensional projections of the original high-dimensional data while trying to minimize distortion. We compare SANJAY to the popular multidimensional scaling (MDS) approach for visualization of small data sets drawn from a representative set of benchmarks, and our experiments show that SANJAY produces distortions that are 1.44 to 4.15 times smaller than those caused due to MDS. Our experimental results show that SANJAY also outperforms the Random Projections technique in terms of the distortions in the projections. CONCLUSIONS: We describe a new algorithmic technique that uses a symbolic decision procedure to automatically synthesize low-dimensional projections of flow cytometry data that typically have a high number of dimensions. Our algorithm is the first application, to our knowledge, of using automated theorem proving for automatically generating highly-accurate, low-dimensional visualizations of high-dimensional data.

Simultaneous hepatosplenic T-cell lymphoma and myelofibrosis.

Hepatosplenic T-cell lymphoma (HSTL) is a rare T-cell neoplasm of the lymphoid system. This type of lymphoma is characterized by sinusoidal infiltration of spleen, liver, bone marrow and lymph nodes by neoplastic lymphocytes. Here, we discuss a patient who had a left axillary lymph node biopsy with characteristic histological and immunohistochemical features of HSTL. In addition, infiltrating neoplastic T-cells and simultaneous characteristic features of myelofibrosis (MF) were also present in the bone marrow biopsy specimen. In contrast to secondary MF, primary MF is a progressive disease and may significantly affect the prognosis of coexisting HSTL. There are few reports in the literature talking about mild bone marrow fibrosis in association with T cell lymphoma, however marked increase in bone marrow fibrosis and HSTL never being reported. This case is shedding light on HSTL and marked increase in bone marrow fibrosis.

Characterization of p38 MAPK isoforms for drug resistance study using systems biology approach.

MOTIVATION: p38 mitogen-activated protein kinase activation plays an important role in resistance to chemotherapeutic cytotoxic drugs in treating multiple myeloma (MM). However, how the p38 mitogen-activated protein kinase signaling pathway is involved in drug resistance, in particular the roles that the various p38 isoforms play, remains largely unknown. METHOD: To explore the underlying mechanisms, we developed a novel systems biology approach by integrating liquid chromatography-mass spectrometry and reverse phase protein array data from human MM cell lines with computational pathway models in which the unknown parameters were inferred using a proposed novel algorithm called modularized factor graph. RESULTS: New mechanisms predicted by our models suggest that combined activation of various p38 isoforms may result in drug resistance in MM via regulating the related pathways including extracellular signal-regulated kinase (ERK) pathway and NFкB pathway. ERK pathway regulating cell growth is synergistically regulated by p38δ isoform, whereas nuclear factor kappa B (NFкB) pathway regulating cell apoptosis is synergistically regulated by p38α isoform. This finding that p38δ isoform promotes the phosphorylation of ERK1/2 in MM cells treated with bortezomib was validated by western blotting. Based on the predicted mechanisms, we further screened drug combinations in silico and found that a promising drug combination targeting ERK1/2 and NFκB might reduce the effects of drug resistance in MM cells. This study provides a framework of a systems biology approach to studying drug resistance and drug combination selection. AVAILABILITY AND IMPLEMENTATION: RPPA experimental Data and Matlab source codes of modularized factor graph for parameter estimation are freely available online at http://ctsb.is.wfubmc.edu/publications/modularized-factor-graph.php.

Modeling cell-cell interactions in regulating multiple myeloma initiating cell fate.

Cancer initiating cells have been documented in multiple myeloma and believed to be a key factor that initiates and drives tumor growth, differentiation,metastasis, and recurrence of the diseases. Although myeloma initiating cells (MICs) are likely to share many properties of normal stem cells, the underlying mechanisms regulating the fate of MICs are largely unknown. Studies designed to explore such communication are urgently needed to enhance our ability to predict the fate decisions of ICs (self-renewal, differentiation, and proliferation). In this study, we developed a novel system to understand the intercellular communication between MICs and their niche by seamlessly integrating experimental data and mathematical model. We first designed dynamic cell culture experiments and collected three types of cells (side population cells, progenitor cells, and mature myeloma cells) under various cultural conditions with flow cytometry. Then we developed a lineage model with ordinary differential equations by considering secreted factors, self-renewal, differentiation, and other biological functions of those cells, to model the cell­cell interactions among the three cell types. Particle swarm optimization was employed to estimate the model parameters by fitting the experimental data to the lineage model. The theoretical results show that the correlation coefficient analysis can reflect the feedback loops among the three cell types, the intercellular feedback signaling can regulate cell population dynamics, and the culture strategies can decide cell growth. This study provides a basic framework of studying cell­cell interactions in regulating MICs fate.

Cancer stem cells: a review of potential clinical applications.

CONTEXT: Cancer stem cells (CSCs) comprise a minor cell population in a tumor; however, they possess self-renewal capacity and are responsible for tumor recurrence and the emerging issue of tumor resistance. Despite recent advances in the study of pathogenesis and mechanisms of CSC-mediated disease recurrence and multidrug resistance, many questions remain unanswered. OBJECTIVES: To provide an overview of CSC theory and to describe major methods of CSC detection and isolation, with the emphasis on those techniques that are potentially relevant in clinical laboratory practice. Particular attention is given to CSC markers, such as cancer testis antigens, which could become promising targets in the development of immunotherapy in settings of minimal residual disease. DATA SOURCES: The review is based on analysis of peer-reviewed literature cited in PubMed, as well as preliminary results of studies conducted in our laboratory. CONCLUSIONS: Despite a lack of consensus in the scientific community on research methodology, CSCs have demonstrated significant potential as therapeutic targets in the treatment of cancer. Further research of CSC biology and markers will eventually lead to the development of novel therapeutic approaches for targeting these cells to treat resistant and recurrent tumors and minimal residual disease.

Differentiating between Burkitt lymphoma and CD10+ diffuse large B-cell lymphoma: the role of commonly used flow cytometry cell markers and the application of a multiparameter scoring system.

The goal of this study was to evaluate routine flow cytometric (FC) immunophenotypic markers in differentiating between Burkitt lymphoma (BL) and CD10+ diffuse large B-cell lymphoma (DLBCL). We performed retrospective analysis of FC data from 55 patients. We evaluated 9 FC parameters: forward and side scatter (FSC and SSC); mean fluorescent intensity (MFI) for CD20, CD10, CD38, CD79b, CD43, and CD71; and the percentage of neoplastic cells positive for CD71 (%CD71). The FSC; MFIs of CD10, CD43, CD79b, and CD71; and %CD71 cells were significantly different between BL and CD10+ DLBCL (P < .05; Student t test). A 5-point scoring system (FSC, %CD71, and MFIs of CD43, CD79b, and CD71) was devised, and 6 (60%) of 10 BLs scored 3 or greater and 1 (10%) of 10 CD10+ DLBCLs scored 3 (P = .04; χ(2)). Our findings indicate that routine FC parameters can aid in differentiating BL from CD10+ DLBCL.

p38 Mitogen-activated protein kinase and hematologic malignancies.

CONTEXT: p38 mitogen-activated protein kinase (MAPK) signaling has been implicated in responses ranging from apoptosis to cell cycle, induction of expression of cytokine genes, and differentiation. This plethora of activators conveys the complexity of the p38 pathway. This complexity is further complicated by the observation that the downstream effects of p38 MAPK activation may be different depending on types of stimuli, cell types, and various p38 MAPK isoforms involved. OBJECTIVE: This review focuses on the recent advancement of the p38 MAPK isoforms as well as the roles of p38 MAPK in hematologic malignancies. DATA SOURCES: Review of pertinent published literature and work in our laboratory. CONCLUSIONS: In some hematologic malignancies, activation of p38 plays a key role in promoting or inhibiting proliferation and also in increasing resistance to chemotherapeutic agents. The importance of different p38 isoforms in various cellular functions has been acknowledged recently. Further understanding of these isoforms will allow the design of more specific inhibitors to target particular isoforms to maximize the treatment effect and minimize the side effects for treating hematopoietic malignancies.

Intraocular lymphoma: diagnostic approach and immunophenotypic findings in vitrectomy specimens.

CONTEXT: Diagnosis and classification of primary intraocular lymphoma can be challenging because of the sparse cellularity of the vitreous specimens. OBJECTIVE: To classify and clinically correlate intraocular lymphoma according to the World Health Organization (WHO) classification by using vitrectomy specimens. DESIGN: Clinical history, cytologic preparations, flow cytometry reports, and outcome of 16 patients diagnosed with intraocular lymphoma were reviewed. RESULTS: The study group included 10 women and 6 men. The mean age of the patients was 63 years (range, 19-79 years). Eleven patients had central nervous system involvement and 6 patients had systemic involvement. All cases were adequately diagnosed and classified according to the WHO classification by using combination of cytologic preparations and 4-color flow cytometry with a limited panel of antibodies to CD19, CD20, CD5, CD10, and kappa and lambda light chains. The cases included 9 primary diffuse large B-cell lymphomas of the CNS type; 2 diffuse large B-cell lymphomas, not otherwise specified; 1 extranodal, low-grade, marginal zone lymphoma of mucosa-associated lymphoid tissue (MALT); 1 precursor B-lymphoblastic lymphoma; and 3 peripheral T-cell lymphomas, not otherwise specified. Of note, all 11 cases of diffuse large B-cell lymphoma were CD10-. All the patients received systemic chemotherapy and radiation therapy. Only 4 patients were free of disease at last follow-up (range, 18 months to 8 years), with severe visual loss. CONCLUSIONS: Intraocular lymphoma cases can be adequately classified according to the WHO classification. Diffuse large B-cell lymphoma, CD10- and most likely of non-germinal center B-cell-like subgroup, is the most common subtype of non-Hodgkin lymphoma in this site, in contrast to ocular adnexal lymphoma for which MALT lymphoma is the most common subtype.

Polymorphisms in TGFbeta and TNFalpha are associated with the myelodysplastic syndrome phenotype.

CONTEXT: Myelodysplastic syndromes (MDSs) are characterized by ineffective hematopoiesis, excessive apoptosis, and the aberrant expression of a number of cytokines. The genes encoding these cytokines are significantly polymorphic. It is unknown whether these cytokine polymorphisms are associated with, and may therefore be playing a role in the pathogenesis of, MDS. OBJECTIVE: To determine if certain polymorphisms in the tumor necrosis factor alpha (TNF-alpha) and transforming growth factor beta (TGF-beta) cytokines are overrepresented in a cohort of patients with MDSs. DESIGN: DNA was isolated from the peripheral blood or bone marrow aspirate of 21 patients with MDS. The genotypes for 4 different polymorphisms, 2 in TNFalpha and 2 in TGFbeta1, were determined using single-specific-primer polymerase chain reaction. The allele and genotype frequencies were compared with similar populations in the National Cancer Institute SNP500 database. RESULTS: In our MDS population, the -308A/A genotype of the TNFalpha gene and the TGFbeta1 allele +29T and genotype +29T/T, each associated with higher levels of expression, were overrepresented in our MDS population. CONCLUSIONS: Polymorphisms associated with increased expression in the cytokines TNFalpha and TGFbeta1 are overrepresented in the MDS population suggesting that increased TNF-alpha and TGF-beta1 activity may contribute to the susceptibility and/or pathogenesis of MDS. Further studies with larger sample sizes are warranted to confirm our observation.

Histiocytic sarcoma - a case with evenly distributed multinucleated giant cells.

Histiocytic sarcoma is an uncommon neoplasm of mature histiocytes with a poor clinical outcome. We report a case of a true histiocytic sarcoma with prominent and evenly distributed multinucleated giant cells that mimics a giant cell tumor of soft tissue. The tumor was located between the appendix, right ovary, and the terminal ileum with severe adhesion. The liver and spleen were not enlarged. Grossly, the tumor appeared grayish white, solid, and soft. Microscopically, polygonal mononuclear tumor cells aggregated to form somewhat epithelioid nests, which occasionally showed coagulative necrosis. Prominent and evenly scattered giant cells were present in all sections. In addition, tumor cell infiltration was noted in regional lymph nodes. The tumor cells were positive for lysozyme, CD68, CD163, and negative for T- and B-cell lineage markers, follicular dendritic cell, megakaryocytic, epithelial, muscular, and melanocytic markers, CD1a and CD30. This case posed great difficulty in clinical and pathological diagnoses. Gross pictures, microscopic findings, and extensive immunostains are important for the differential diagnosis.

Chronic idiopathic myelofibrosis: clinicopathologic features, pathogenesis, and prognosis.

CONTEXT: Chronic idiopathic myelofibrosis (CIMF) is a clonal myeloproliferative disease characterized by panmyelosis with intact maturation, progressive bone marrow fibrosis, and multiorgan extramedullary hematopoiesis. OBJECTIVE: This review article aims to summarize the recent updates regarding the clinicopathologic features, molecular pathogenesis, cytogenetic abnormalities, diagnostic criteria, new diagnostic ancillary tests, and prognostic factors of CIMF. DATA SOURCES: Important relevant articles indexed in PubMed/MEDLINE (National Library of Medicine) through the end of 2005 and referenced medical texts. CONCLUSIONS: Because CIMF has a variety of clinical presentations, diagnosis may be challenging; the prefibrotic stage of CIMF has always been a challenging disease for pathologists to diagnose accurately. The recently proposed European Clinical and Pathological criteria can be helpful in the diagnosis of CIMF, especially in its prefibrotic stage. The enumeration of CD34-positive cells in the peripheral blood and the presence of circulating endothelial progenitor cells are the new important ancillary tests for the diagnosis of a small subset of patients with CIMF with atypical presentation. The recent discovery of the new mutation affecting the Janus tyrosine kinase 2 (JAK2V617F), more frequently observed in patients with polycythemia vera, is seen in approximately 35% to 57% of patients with CIMF. This mutation can serve as another diagnostic tool. Important factors affecting prognosis in CIMF are anemia, age of the patient, white blood cell count, degree of fibrosis, and number of blasts in the peripheral blood.