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Biomedical subjects

Nancy Berliner

Publications and source records attributed to Nancy Berliner.

18 recordsLinked to original sources
PubMed2006

Effect of Factor V Leiden and prothrombin G20210-->A mutations on thromboembolic risk in the national surgical adjuvant breast and bowel project breast cancer prevention trial.

BACKGROUND: In the National Surgical Adjuvant Breast and Bowel Project's Breast Cancer Prevention Project (BCPT), tamoxifen use was associated with an increased relative risk for venous thromboembolic events, including deep vein thrombosis and pulmonary emboli, compared with placebo. However, the involvement of hypercoagulability factors in this association is unclear. METHODS: To examine possible associations among the risk of venous thromboembolic events, tamoxifen use, and Factor V Leiden (FVL) and prothrombin G20210-->A (PT20210) mutations, which are involved in promoting blood coagulation, we used a nested, matched, case-control (1 : 4) design and compared women in the BCPT who had experienced venous thromboembolic events (n = 76) with women who did not (n = 295). FVL and PT20210 mutations were detected in genomic DNA that was isolated from blood samples collected at trial enrollment. RESULTS: Venous thromboembolic events occurred in 28 women (deep vein thrombosis in 22 and pulmonary emboli in six) who were taking placebo and in 53 women (deep vein thrombosis in 35 and pulmonary emboli in 18) who were taking tamoxifen (relative risk = 1.90, 95% confidence interval = 1.18 to 3.12). Excessive risk for venous thromboembolic events was observed only in the first 36 months of therapy. There were no differences in age, smoking, and race between the groups, but women with venous thromboembolic events had a higher body mass index than women without (mean +/- standard deviation, 30 kg/m(2) +/- 7.7 versus 27.1 +/- 5.6; P<.001). FVL and/or PT20210 mutations were found in nine women (four on tamoxifen and five on placebo) with venous thromboembolic events and in 20 control subjects (nine on tamoxifen and 11 on placebo). No associations were found between risk of venous thromboembolic events and mutation status in either treatment group. CONCLUSIONS: Venous thromboembolic disease in the BCPT women is associated with tamoxifen use and body mass index, but not with FVL and PT20210 mutations. Screening women at risk for breast cancer for FVL and/or PT20210 appears to offer no benefit in determining the risk of tamoxifen-associated thromboembolic events.

Adenine↗
PubMed2006

Treatment-related myelodysplasia/AML in a patient with a history of breast cancer and an oligodendroglioma treated with temozolomide: case study and review of the literature.

The emergence of temozolomide as an effective alkylating agent with little acute toxicity or cumulative myelosuppression has led to protracted courses of chemotherapy for many patients with gliomas. Secondary, or treatment-related, myelodysplasia (t-MDS) and acute myelogenous leukemia (t-AML) are life-threatening complications of alkylating chemotherapy and have been reported in patients with primary brain tumors. We describe a case of temozolomide-related t-MDS/AML and discuss the clinical features of this condition. Administration of an alkylating agent in patient populations with long median survivals must be undertaken with an understanding of the potential for this treatment complication.

Aged↗
PubMed2006

Sustained remission from angioimmunoblastic T-cell lymphoma induced by alemtuzumab.

BACKGROUND: A 73-year-old woman presented with acute lower back pain, fever, chills and arthralgias. She had previously had a positive protein derivative test with a negative chest X-ray; her medical history was also remarkable for a mitral valve prolapse. Initial symptoms resolved spontaneously without therapy, but fever recurred with associated arthralgias, myalgias, diffuse and worsening lymphadenopathy, splenomegaly, and bilateral pulmonary infiltrates. INVESTIGATIONS: Physical examination, blood and urine cultures, MRI of the spine, echocardiogram, extensive serologies, serum and urine protein electrophoresis, immunofixation electrophoresis, bone-marrow aspiration and biopsy with flow cytometry, cytogenetics, and gene rearrangement studies, CT scan of the chest, abdomen and pelvis, whole-body PET, and lymph-node biopsy for histological examination, immunohistochemistry, and gene rearrangement studies. DIAGNOSIS: Angioimmunoblastic T-cell lymphoma. MANAGEMENT: Steroids (prednisone, methylprednisolone), levofloxacin, isoniazid with pyridoxine, ciclosporin A, methotrexate, alemtuzumab, broad-spectrum antibiotics, Pneumocystis carinii prophylaxis, vancomycin, and clindamycin.

Aged↗
PubMed2006

p120 nucleolar-proliferating antigen is a direct target of G-CSF signaling during myeloid differentiation.

Granulocyte-colony stimulating factor (G-CSF) is an essential cytokine, which contributes to proliferation and differentiation of granulocyte precursor cells in the bone marrow. Despite recent progress in understanding G-CSF signaling events, the mechanisms that underlie the distinct spectrum of biological functions attributed to G-CSF-mediated gene expression remain unclear. Previous studies have identified a number of genes, which are up-regulated in G-CSF-stimulated myeloid precursor cells. In this study, we sought to identify additional target genes of G-CSF-mediated proliferation and/or differentiation. cDNA representational difference analysis was used with the 32Dcl3 cell line as a model system to isolate genes, which are up-regulated in an immediate-early manner upon G-CSF stimualtion. We isolated p120 nucleolar-proliferation antigen (NOL1), a highly conserved, nucleolar-specific, RNA-binding protein of unknown function, and confirmed its expression by Northern blot analysis in 4-h, G-CSF-induced 32Dcl3 cells. Isolation of a mouse p120 genomic clone revealed the presence of a signal tranducer and activator of transcription (STAT)-binding site in the first intron of the gene. We demonstrate the importance of STAT3 and STAT5 in mediating the G-CSF response with respect to p120 expression by transient transfection analysis, oligonucleotide pull-down assays, and the loss of p120 expression in the bone marrow of mice lacking normal STAT3 signaling. In addition, overexpression of p120 in G-CSF-induced 32D cells revealed normal, morphologic maturation and growth characteristics but loss of lactoferrin expression, a marker of normal neutrophil maturation, suggesting that inappropriate expression of the p120 gene can result in aberrant neutrophil maturation.

Animals↗
PubMed2005

Heterogeneity of functional responses in differentiated myeloid cell lines reveals EPRO cells as a valid model of murine neutrophil functional activation.

Mature neutrophils display multiple functional responses upon activation that include chemotaxis, adhesion to and transmigration across endothelial cells, phagocytosis, and pathogen destruction via potent microbicidal enzymes and reactive oxygen species. We are using myeloid cell line models to investigate the signaling pathways that govern neutrophil functional activation. To facilitate these studies, we have performed a direct comparison of functional responses of human and murine myeloid cell line models upon neutrophil differentiation. Our results show that EPRO cells, promyelocytes that undergo complete neutrophil maturation, demonstrate a full spectrum of functional responses, including respiratory burst, chemotaxis toward two murine chemokines, and phagocytosis. We also extend previous studies of granulocyte-colony stimulating factor-induced 32Dcl3 cells, showing they demonstrate chemotaxis and phogocytosis but completely lack a respiratory burst as a result of the absent expression of a critical oxidase subunit, gp91(phox). Induced human leukemic NB4 and HL-60 cells display a respiratory burst and phagocytosis but have defective chemotaxis to multiple chemoattractants. We also tested each cell line for the ability to up-regulate cell-surface membrane-activated complex-1 (Mac-1) expression upon activation, a response mediating neutrophil adhesion and a surrogate marker for degranulation. We show that EPRO cells, but not 32Dcl3 or NB4, significantly increase Mac-1 surface expression upon functional activation. Together, these data show that EPRO and MPRO cells demonstrate complete, functional activation upon neutrophil differentiation, suggesting these promyelocytic models accurately reflect the functional capacity of mature murine neutrophils.

Animals↗
PubMed2005

Human neutrophil collagenase expression is C/EBP-dependent during myeloid development.

OBJECTIVE: Human neutrophil collagenase (HNC) is one of several secondary granule proteins (SGP) expressed late in the myeloid maturation pathway. SGPs are encoded by unlinked and functionally diverse genes that are hypothesized to be coordinately regulated at the transcriptional level and demonstrate uniform dysregulation in leukemic cells. In support of the hypothesis that tissue and stage-specific expression of SGP genes is regulated by shared factor(s), we sought to identify factors responsible for positive regulation of the SGP genes. METHODS: Using 5' deletion analysis, we identified a minimal HNC promoter located within the first 193 bp upstream of the transcription start site. Three CCAAT enhancer binding protein (C/EBP) sites were identified within this region and their functional importance was confirmed by mutational analysis, gel retardation, and oligonucleotide pulldown assays. Using chromatin immunoprecipitation (ChIP), we demonstrated that C/EBPalpha binds to the SGP gene promoters lactoferrin and HNC in nonexpressing cells. Upon induction of maturation, C/EBPalpha binds to these promoters and this binding correlates with the expression of both SGP genes. CONCLUSION: We conclude that in the later stages of myeloid development, SGP genes are coordinately upregulated, and that members of the C/EBP family of transcription factors, in particular C/EBPalpha and C/EBPepsilon, play specific and unique roles in upregulating their expression.

Animals↗
PubMed2005

Modulation of NB4 promyelocytic leukemic cell machinery by Anaplasma phagocytophilum.

Anaplasma phagocytophilum is a gram-negative obligate intracellular bacterium that persists within neutrophils. We assessed the impact of A. phagocytophilum infection in NB4 promyelocytic leukemic cells using high-density oligoarray, two-dimensional differential gel electrophoresis and liquid chromatography-mass spectrometry. Our Affymetrix data revealed that A. phagocytophilum altered the expression of transcription factors, cell adhesion molecules, signal transduction genes, and proinflammatory cytokines. However, the expression of Toll-like receptors, MYD88, RNF36, IRF3, and TBK1 and inhibitors of the NF-kappaB gene was not altered. A. phagocytophilum infection also altered the apoptotic program of NB4 cells and resulted in increased transcription of antiapoptotic genes (MCL1 and BFL1). The transcription and translation of iron-metabolism genes (light polypeptide ferritin chain, transferrin, and the transferrin receptor) were significantly altered, suggesting a possible link between A. phagocytophilum infection and iron metabolism. Our study clearly demonstrates multifactorial effects of A. phagocytophilum infection on NB4 promyelocytic leukemic cell machinery.

Anaplasma phagocytophilum↗
PubMed2005

Anaplasma phagocytophilum modulates gp91phox gene expression through altered interferon regulatory factor 1 and PU.1 levels and binding of CCAAT displacement protein.

Infection of neutrophil precursors with Anaplasma phagocytophilum, the causative agent of human granulocytic ehrlichiosis, results in downregulation of the gp91(phox) gene, a key component of NADPH oxidase. We now show that repression of gp91(phox) gene transcription is associated with reduced expression of interferon regulatory factor 1 (IRF-1) and PU.1 in nuclear extracts of A. phagocytophilum-infected cells. Loss of PU.1 and IRF-1 correlated with increased binding of the repressor, CCAAT displacement protein (CDP), to the promoter of the gp91(phox) gene. Reduced protein expression of IRF-1 was observed with or without gamma interferon (IFN-gamma) stimulation, and the defect in IFN-gamma signaling was associated with diminished binding of phosphorylated Stat1 to the Stat1 binding element of the IRF-1 promoter. The diminished levels of activator proteins and enhanced binding of CDP account for the transcriptional inhibition of the gp91(phox) gene during A. phagocytophilum infection, providing evidence of the first molecular mechanism that a pathogen uses to alter the regulation of genes that contribute to an effective respiratory burst.

Anaplasma phagocytophilum↗
PubMed2005

Early transcriptional response of human neutrophils to Anaplasma phagocytophilum infection.

Anaplasma phagocytophilum, an unusual obligate intracellular pathogen that persists within neutrophils, causes human anaplasmosis (previously known as human granulocytic ehrlichiosis). To study the effects of this pathogen on the transcriptional profile of its host cell, we performed a comprehensive DNA microarray analysis of the early (4-h) transcriptional response of human neutrophils to A. phagocytophilum infection. A. phagocytophilum infection resulted in the up- and down-regulation of 177 and 67 neutrophil genes, respectively. These data were verified by quantitative reverse transcription-PCR of selected genes. Notably, the up-regulation of many antiapoptotic genes, including the BCL2A1, BIRC3, and CFLAR genes, and the down-regulation of the proapoptotic TNFSF10 gene were observed. Genes involved in inflammation, innate immunity, cytoskeletal remodeling, and vesicular transport also exhibited differential expression. Vascular endothelial growth factor was also induced. These data suggest that A. phagocytophilum may alter selected host pathways in order to facilitate its survival within human neutrophils. To gain further insight into the bacterium's influence on host cell gene expression, this report presents a detailed comparative analysis of our data and other gene expression profiling studies of A. phagocytophilum-infected neutrophils and promyelocytic cell lines.

Anaplasma phagocytophilum↗
PubMed2004

Congenital and acquired neutropenia.

Our understanding of the pathogenesis of congenital and acquired neutropenia is rapidly evolving. New ground-breaking observations have identified the genes responsible for many of the congenital neutropenia syndromes and are also providing new insights into normal neutrophil commitment and differentiation. Acquired neutropenia remains a poorly understood syndrome, although new insights into its pathogenesis are also emerging, especially with regard to subsets of immune neutropenia. In Section I, Dr. Marshall Horwitz reviews the current understanding of the genetic basis, molecular pathology, and approaches to treatment of congenital neutropenia and cyclic hematopoiesis. Mutations in the ELA2 gene, which encodes for neutrophil elastase, cause cyclic hematopoiesis. ELA2 mutations are also the most common cause of congenital neutropenia, where their presence may equate with a more severe clinical course and higher frequency of leukemic progression. Emerging evidence indicates interrelatedness with Hermansky Pudlak syndrome and other disorders of neutrophil and platelet granules. In Section II, Dr. Nancy Berliner presents an overview of the clinical approach to the evaluation and treatment of acquired neutropenia. This includes a review of the pathogenesis of primary and secondary immune neutropenia, drug-induced neutropenia, and non-immune chronic idiopathic neutropenia of adults. Studies used to evaluate patients for potential immune neutropenia are reviewed. Management issues, especially the use of granulocyte colony-stimulating factor (G-CSF), are discussed. In Section III, Dr. Thomas Loughran, Jr., reviews the pathogenesis and clinical manifestations of large granular lymphocyte (LGL) leukemia. Possible mechanisms of neutropenia are discussed. In particular, discussion focuses on the relationship between LGL leukemia, rheumatoid disease, and Felty's syndrome, and the complex interplay of defects in neutrophil production, distribution, destruction, and apoptosis that underly the development of neutropenia in those syndromes.

Antigen-Antibody Complex↗
PubMed2004

T-cell large granular lymphocyte leukemia and related disorders.

T-cell large granular lymphocyte (LGL) leukemia is a clonal proliferation of cytotoxic T cells, which causes neutropenia, anemia, and/or thrombocytopenia. This condition is often associated with autoimmune disorders, especially rheumatoid arthritis, and other lymphoproliferative disorders. The diagnosis is suggested by flow cytometry demonstrating an expansion of CD8(+)CD57(+) T cells and is confirmed by T-cell receptor gene rearrangement studies. Mounting evidence suggests that LGL leukemia is a disorder of dysregulation of apoptosis through abnormalities in the Fas/Fas ligand pathway. In most patients, this is an indolent disorder, and significant improvement of cytopenias can be achieved with immunosuppressive agents such as steroids, methotrexate, cyclophosphamide, and cyclosporin A. This review provides a concise, up-to-date summary of LGL leukemia and the related, more aggressive, malignancies of cytotoxic T cells and natural killer cells.

Aged↗
PubMed2004

Molecular diagnostics of malignant disorders.

Expansion of our understanding of the molecular basis of cancer has enabled us to apply molecular techniques to categorize malignancies into more uniform, informative groups. In this review we describe the basic molecular techniques used for this purpose, including Southern blotting, polymerase chain reaction (PCR) and quantitative PCR, fluorescence in situ hybridization, DNA microarrays, and proteomics. The main applications of these techniques in the modern management of acute leukemia, chronic myeloid leukemia, lymphomas, and breast cancer are summarized.

Blotting, Southern↗
PubMed2003

G-CSF signaling can differentiate promyelocytes expressing a defective retinoic acid receptor: evidence for divergent pathways regulating neutrophil differentiation.

Several lines of investigation suggest that granulocyte colony-stimulating factor (G-CSF) augments all-trans retinoic acid (ATRA)-induced neutrophil differentiation in acute promyelocytic leukemia (APL). We sought to characterize the relationship between G-CSF- and ATRA-mediated neutrophil differentiation. We established a G-CSF receptor-transduced promyelocytic cell line, EPRO-Gr, derived from the granulocyte-macrophage colony-stimulating factor (GM-CSF)-dependent EPRO cell line harboring a dominant-negative retinoic acid receptor alpha (RARalpha). In EPRO-Gr, neutrophil differentiation occurs either in GM-CSF upon addition of ATRA or upon induction with G-CSF alone. Transient transfection of EPRO-Gr cells with a RARE-containing reporter plasmid demonstrates increased activity in the presence of ATRA, but not G-CSF, while STAT3 phosphorylation occurs only in response to G-CSF. This suggests that ATRA-mediated differentiation of EPRO-Gr cells occurs via a RARE-dependent, STAT3-independent pathway, while G-CSF-mediated differentiation occurs via a RARE-independent, STAT3-dependent pathway. ATRA and G-CSF thus regulate differentiation by divergent pathways. We characterized these pathways in the APL cell line, NB4. ATRA induction of NB4 cells resulted in morphologic differentiation and up-regulation of C/EBPepsilon and G-CSFR, but not in STAT3 phosphorylation. The addition of G-CSF with ATRA during NB4 induction resulted in STAT3 phosphorylation but did not enhance differentiation. These results may elucidate how G-CSF and ATRA affect the differentiation of primary and ATRA-resistant APL cells.

Animals↗
PubMed2003

Chromatin immunoprecipitation (ChIP) studies indicate a role for CCAAT enhancer binding proteins alpha and epsilon (C/EBP alpha and C/EBP epsilon ) and CDP/cut in myeloid maturation-induced lactoferrin gene expression.

In vitro models of granulopoiesis involving the inducible expression of either CCAAT enhancer binding protein alpha (C/EBP alpha) or C/EBP epsilon in myeloid cells have been shown to lead to the induction of a granulocytic maturation program accompanied by the expression of myeloid-specific genes. Since members of the C/EBP family of transcription factors recognize and bind to similar DNA-binding motifs, it has been difficult to elucidate the specific role of each of the C/EBP family members in eliciting myeloid gene expression. In order to address this issue, we focused on the expression of the lactoferrin (LF) gene. LF expression is transcriptionally regulated in a C/EBP-dependent manner in myeloid cells. Using chromatin immunoprecipitation (ChIP) analysis we demonstrate that C/EBP alpha binds to the LF promoter in nonexpressing cells. Upon induction of maturation, C/EBP epsilon binds to the LF promoter, which correlates with LF expression. Lack of LF expression in the acute promyelocytic leukemia cell line NB4, which harbors the t(15;17) translocation, cannot be correlated with aberrant binding at the C/EBP site in the LF promoter. It is, however, associated with the persistent binding of the silencer CCAAT displacement protein (CDP/cut) to the LF promoter in these cells. We conclude that C/EBP alpha, C/EBP epsilon, and CDP/cut all play definitive roles in regulating late gene expression during normal myeloid development.

CCAAT-Enhancer-Binding Protein-alpha↗
PubMed2002

Genomic and proteomic analysis of the myeloid differentiation program: global analysis of gene expression during induced differentiation in the MPRO cell line.

We have used an approach using 2-dimensional gel electrophoresis with mass spectrometry analysis combined with oligonucleotide chip hybridization for a comprehensive and quantitative study of the temporal patterns of protein and mRNA expression during myeloid development in the MPRO murine cell line. This global analysis detected 123 known proteins and 29 "new" proteins out of 220 protein spots identified by tandem mass spectroscopy, including proteins in 12 functional categories such as transcription factors and cytokines. Bioinformatic analysis of these proteins revealed clusters with functional importance to myeloid differentiation. Previous analyses have found that for a substantial number of genes the absolute amount of protein in the cell is not strongly correlated to the amount of mRNA. These conclusions were based on simultaneous measurement of mRNA and protein at just a single time point. Here, however, we are able to investigate the relationship between mRNA and protein in terms of simultaneous changes in their levels over multiple time points. This is the first time such a relationship has been studied, and we find that it gives a much stronger correlation, consistent with the hypothesis that a substantial proportion of protein change is a consequence of changed mRNA levels, rather than posttranscriptional effects. Cycloheximide inhibition also showed that most of the proteins detected by gel electrophoresis were relatively stable. Specific investigation of transcription factor mRNA representation showed considerable similarity to those of mature human neutrophils and highlighted several transcription factors and other functional nuclear proteins whose mRNA levels change prominently during MPRO differentiation but which have not been investigated previously in the context of myeloid development. Data are available online at http://bioinfo.mbb.yale.edu/expression/myelopoiesis.

Animals↗
PubMed2002

Development of a murine hematopoietic progenitor complementary DNA microarray using a subtracted complementary DNA library.

With the goal of creating a resource for in-depth study of myelopoiesis, we have executed a 2-pronged strategy to obtain a complementary DNA (cDNA) clone set enriched in hematopoietic genes. One aspect is a library subtraction to enrich for underrepresented transcripts present at early stages of hematopoiesis. For this, a hematopoietic cDNA library from primary murine bone marrow cells enriched for primitive progenitors was used as tester. The subtraction used 10 000 known genes and expressed sequence tags (ESTs) as driver. The 2304 randomly picked clones from the subtracted cDNA libraries represent 1255 distinct genes, of which 622 (50%) are named genes, 386 (30%) match uncharacterized ESTs, and 247 (20%) are novel. The second aspect of our strategy was to complement this subtracted library with genes known to be involved in myeloid cell differentiation and function. The resulting cDNAs were arrayed on polylysine-coated glass slides. The microarrays were used to analyze gene expression in primary and cultured murine bone marrow-derived progenitors. We found expression of various types of genes, including regulatory cytokines and their receptors, signal transduction genes, and transcription factors. To assess gene expression during myeloid differentiation, we examined patterns of change during induced differentiation of EML cells. Several hundred of the genes underwent fluctuations in expression level during myeloid cell differentiation. The complete database, accessible on the World Wide Web at http://yale130132115135.med.yale.edu/, allows for retrieval of information regarding these genes. Our microarray allows for genomewide expression analysis of myeloid stem cells, which will help in defining the regulatory mechanisms of stem cell differentiation.

Animals↗