Implication of HIF-1α but Not HIF-2α in the Hypoxic Response of Human Hematopoietic Progenitors.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 4154-4154
Author(s):  
Yanyan Zhang ◽  
Adlen Foudi ◽  
Magali Berthebaud ◽  
Dorothee Buet ◽  
Peggy Jarrier ◽  
...  
Keyword(s):  
Mrna Expression ◽  
Cell Lines ◽  
Hematopoietic Cells ◽  
Receptor Expression ◽  
Metabolic Adaptation ◽  
Protein Accumulation ◽  
Mrna Levels ◽  
Hematopoietic Progenitors ◽  
Cd34 Cells ◽  
Cxcr4 Mrna

Abstract Maturing hematopoietic cells are exposed to hypoxia as they develop and migrate within the bone marrow microenvironment. Previous studies using non hematopoietic cell lines and monocytes showed that CXCR4 is strongly induced by hypoxia but little is known on the regulation of CXCR4 by hypoxia in the other hematopoietic cells and during hematopoietic development. We analyzed the expression and regulation of hypoxia-inducible transcription factor-1a (HIF1a) and 2a (HIF2a), the master regulators of metabolic adaptation to hypoxia, during hematopoiesis. Real time quantitative RT-PCR showed that HIF-1a mRNA was present on all the non hematopoietic and hematopoietic cells lines including HL-60, HEL, TF1, K562, KG1, U937, Jurkat and Mo7e. In contrast, HIF-2a mRNA expression was variable among the cell lines and was detected only at very low level in some cells such as KG1, Jurkat and HEL. Hypoxia exposure rapidly induced VEGF mRNA expression in the cells that expressed HIF-1a mRNA and exhibited HIF-1a protein accumulation. Interestingly, CXCR4 induction was observed only in the cells that exhibit significant expression of HIF-2a mRNA and HIF-2a protein accumulation. A strong correlation between HIF-2a mRNA levels and the induction of CXCR4 mRNA expression by hypoxia was found. Human CD34+ cells also expressed high levels of HIF-1a mRNA, whereas HIF-2a mRNA was barely detected. Interestingly, as observed for several myeloid cell lines, CD34+ cells exhibited a strong induction of VEGF expression in response to hypoxia and hypoxia mimetic agents cobalt chloride and desferrrioxamine whereas CXCR4 receptor expression was not induced suggesting that CXCR4 mRNA induction is related to the expression of HIF-2a. Altogether these data indicated that the hypoxic responses of human hematopoietic progenitors are independent of HIF-2a. Moreover, they establish that CXCR4 regulation by hypoxia is linked to HIF-2a protein expression.

RASSF2 Functions As a Tumor Suppressor in t(8;21) AML Via Promotion of Apoptosis

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 3590-3590
Author(s):  
Samuel A Stoner ◽  
Russell Dekelver ◽  
Miao-Chia Lo ◽  
Dong-Er Zhang
Keyword(s):  
Gene Expression ◽  
Tumor Suppressor ◽  
Cell Lines ◽  
Hematopoietic Cells ◽  
Leukemia Cell ◽  
Leukemia Cells ◽  
Mrna Levels ◽  
Human Cd34 ◽  
Leukemia Cell Lines ◽  
Cd34 Cells

Abstract The t(8;21) chromosomal translocation is one of the most common chromosomal translocations associated with acute myeloid leukemia (AML), found in approximately 12% of de novo AML cases. The majority of these cases are classified as FAB-subtype M2 AML. The t(8;21) results in the stable fusion of the AML1 (RUNX1) and ETO (RUNX1T1) genes. The AML1-ETO fusion protein is composed of the N-terminal portion of AML1, which includes the DNA-binding Runt-homology domain, and nearly the full-length ETO protein. The primary accepted mechanism by which AML1-ETO promotes leukemia development is through the aberrant recruitment of transcriptional repression/activation complexes to normal AML1 target genes. Therefore, the identification of individual genes or biological pathways that are specifically disrupted in the presence of AML1-ETO will provide further molecular insight into the pathogenesis of t(8;21) AML and lead to the possibility for improved treatment for these patients. We identified RASSF2 as a gene that is specifically downregulated in (2-4 fold) in total bone marrow of t(8;21) patients compared to non-t(8;21) FAB-subtype M2 AML patients by analyzing publicly available gene expression datasets. Similarly, using a mouse model of t(8;21) AML we found Rassf2 mRNA levels to be nearly 30-fold lower in t(8;21) leukemia cells compared to wild-type Lin-Sca-cKit+ (LK) myeloid progenitors. Gene expression analysis by RT-qPCR in leukemia cell lines confirmed that RASSF2 mRNA levels are significantly downregulated (8-10-fold) in both Kasumi-1 and SKNO-1 t(8;21) cell lines as compared to a similar non-t(8;21) HL-60 cell line and to primary human CD34+ control cells. In addition, expression of AML1-ETO in HL-60 or CD34+ cells results in a decrease in RASSF2 mRNA expression, which further suggests that RASSF2 is a target for regulation by AML1-ETO. Assessment of published ChIP-seq data shows that AML1-ETO binds the RASSF2 gene locus at two distinct regions in both primary t(8;21) AML patient samples and in the Kasumi-1 and SKNO-1 cell lines. These regions are similarly bound by several important hematopoietic transcription factors in primary human CD34+ cells, including AML1, ERG, FLI1, and TCF7L2, implicating these two regions as important for the regulation of RASSF2 expression during blood cell differentiation. Overexpression of RASSF2 in human leukemia cell lines using an MSCV-IRES-GFP (MIG) construct revealed that RASSF2 has a strong negative effect on leukemia cell proliferation and viability. The overall percentage of GFP-positive cells in MIG-RASSF2 transduced cells markedly decreased compared to MIG-control transduced cells over a period of 14 days. This effect was primarily due to significantly increased apoptosis in the RASSF2 expressing cell populations. Similarly, we found that expression of RASSF2 significantly inhibits the long-term self-renewal capability of hematopoietic cells transduced with AML1-ETO in a serial replating/colony formation assay. AML1-ETO transduced hematopoietic cells were normally capable of serial replating for more than 6 weeks. However, AML1-ETO transduced cells co-expressing RASSF2 consistently had reduced colony number and lost their ability to replate after 3-4 weeks. This was due to a dramatically increased rate of apoptosis in RASSF2 expressing cells. RASSF2 is reported to be a tumor suppressor that is frequently downregulated at the transcriptional level by hypermethylation in primary tumor samples, but not healthy controls. Here we have identified RASSF2 as a target for repression, and demonstrated its tumor suppressive function in t(8;21) leukemia cells. Further insights into the molecular mechanisms of RASSF2 function in AML will continue to be explored. Disclosures No relevant conflicts of interest to declare.

scholarly journals Expression Profiling of Nuclear Receptors in the NCI60 Cancer Cell Panel Reveals Receptor-Drug and Receptor-Gene Interactions

2010 ◽  
Vol 24 (6) ◽  
pp. 1287-1296 ◽  
Author(s):  
Susan Holbeck ◽  
Jianjun Chang ◽  
Anne M. Best ◽  
Angie L. Bookout ◽  
David J. Mangelsdorf ◽  
...  
Keyword(s):  
Drug Interactions ◽  
Cell Lines ◽  
Nuclear Receptors ◽  
Cancer Cell ◽  
Expression Profiling ◽  
Expression Profiles ◽  
Receptor Gene ◽  
Receptor Expression ◽  
Cancer Drug ◽  
Mrna Levels

Abstract We profiled the expression of the 48 human nuclear receptors (NRs) by quantitative RT-PCR in 51 human cancer cell lines of the NCI60 collection derived from nine different tissues. NR mRNA expression accurately classified melanoma, colon, and renal cancers, whereas lung, breast, prostate, central nervous system, and leukemia cell lines exhibited heterogeneous receptor expression. Importantly, receptor mRNA levels faithfully predicted the growth-inhibitory qualities of receptor ligands in nonendocrine tumors. Correlation analysis using NR expression profiles and drug response information across the cell line panel uncovered a number of new potential receptor-drug interactions, suggesting that in these cases, individual receptor levels may predict response to chemotherapeutic interventions. Similarly, by cross-comparing receptor levels within our expression dataset and relating these profiles to existing microarray gene expression data, we defined interactions among receptors and between receptors and other genes that can now be mechanistically queried. This work supports the strategy of using NR expression profiling to classify various types of cancer, define NR-drug interactions and receptor-gene networks, predict cancer-drug sensitivity, and identify druggable targets that may be pharmacologically manipulated for potential therapeutic intervention.

scholarly journals Synaptotagmin 13 Is Highly Expressed in Estrogen Receptor-Positive Breast Cancer

2021 ◽  
Vol 28 (5) ◽  
pp. 4080-4092
Author(s):  
Takahiro Ichikawa ◽  
Masahiro Shibata ◽  
Takahiro Inaishi ◽  
Ikumi Soeda ◽  
Mitsuro Kanda ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Estrogen Receptor ◽  
Mrna Expression ◽  
Cell Lines ◽  
Mrna Levels ◽  
Pcr Array ◽  
Array Analysis ◽  
Expression Levels ◽  
Er Positive ◽  
Mrna Expression Levels

Background: Accumulating evidence indicates tumor-promoting roles of synaptotagmin 13 (SYT13) in several cancers; however, no studies have investigated its expression in breast cancer (BC). This study aimed to clarify the significance of SYT13 in BC. Methods: SYT13 mRNA expression levels were evaluated in BC cell lines. Polymerase chain reaction (PCR) array analysis was conducted to determine the correlation between expression levels of SYT13 and other tumor-associated genes. Then, the association of SYT13 expression levels in the clinical BC specimens with patients’ clinicopathological factors was evaluated. These findings were subsequently validated using The Cancer Genome Atlas (TCGA) database. Results: Among 13 BC cell lines, estrogen receptor (ER)-positive cells showed higher SYT13 mRNA levels than ER-negative cells. PCR array analysis revealed positive correlations between SYT13 and several oncogenes predominantly expressed in ER-positive BC, such as estrogen receptor 1, AKT serine/threonine kinase 1, and cyclin-dependent kinases 4. In 165 patients, ER-positive specimens exhibited higher SYT13 mRNA expression levels than ER-negative specimens. The TCGA database analysis confirmed that patients with ER-positive BC expressed higher SYT13 levels than ER-negative patients. Conclusion: This study suggests that SYT13 is highly expressed in ER-positive BC cells and clinical specimens, and there is a positive association of SYT13 with the ER signaling pathways.

scholarly journals FLT3 receptor expression on the surface of normal and malignant human hematopoietic cells

Blood ◽  
1996 ◽  
Vol 88 (9) ◽  
pp. 3383-3390 ◽  
Author(s):  
AM Turner ◽  
NL Lin ◽  
S Issarachai ◽  
SD Lyman ◽  
VC Broudy
Keyword(s):  
Cell Lines ◽  
Myeloid Leukemia ◽  
Hematopoietic Cells ◽  
Leukemia Cell ◽  
Colony Growth ◽  
Receptor Expression ◽  
Flt3 Ligand ◽  
Leukemia Cell Lines ◽  
Kit Receptor ◽  
Myeloid Leukemia Cell

FLT3 ligand is a hematopoietic growth factor that plays a key role in growth of primitive hematopoietic cells. FLT3 receptor mRNA is found in early hematopoietic progenitors and in human myeloid leukemia blasts. Much less is known about the surface expression of FLT3 receptor on human hematopoietic cells. Using human 125I-FLT3 ligand, we have identified and characterized surface FLT3 receptors on normal and malignant human hematopoietic cells and cell lines. Our results showed that surface display of FLT3 receptor was greatest in fresh myeloid leukemia blast cells and myeloid leukemia cell lines. Erythroleukemic and megakaryocytic leukemia cell lines (n = 5) bound little to no 125I-FLT3 ligand. Scatchard analysis of 125I-FLT3 ligand binding data shows that three myeloid leukemia cell lines, ML-1, AML-193, and HL-60, as well as normal human marrow mononuclear cells, exhibit high affinity FLT3 receptors. Crosslinking of 125I-FLT3 ligand to FLT3 receptors on the surface of ML-1 myeloid leukemia cells indicates that the FLT3 ligand. The rates of FLT3 ligand internalization and degradation were determined by binding 125I-FLT3 ligand to ML-1 cells and acid stripping to distinguish surface bound from internalized ligand. Internalized 125I-FLT3 ligand was detected within 5 minutes after binding to ML-1 cells. In addition, we evaluated the effect of FLT3 ligand on megakaryocytic colony growth and nuclear endoreduplication, alone or in the presence of thrombopoietin. FLT3 ligand did not promote colony forming unit megakaryocyte (CFU-Meg) colony growth or megakaryocyte nuclear maturation, nor did FLT3 ligand augment the effects of thrombopoietin on these measures of megakaryopoiesis. These data indicate that the FLT3 receptor shares several characteristics with the c-kit receptor including dimerization and rapid internalization. However, the more restricted cellular distribution of the FLT3 receptor may target the effects of FLT3 ligand to primitive hematopoietic cells and to myeloid and lymphoid progenitor cells, in contrast to the pleiotropic effects of the c-kit receptor ligand, stem cell factor.

scholarly journals Hematopoietic progenitors and interleukin-3-dependent cell lines synthesize histamine in response to calcium ionophore

Blood ◽  
1996 ◽  
Vol 87 (8) ◽  
pp. 3161-3169 ◽  
Author(s):  
M Dy ◽  
A Arnould ◽  
FM Lemoine ◽  
F Machavoine ◽  
H Ziltener ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Mrna Expression ◽  
Cell Lines ◽  
Histidine Decarboxylase ◽  
Calcium Ionophore ◽  
Ionophore A23187 ◽  
Hematopoietic Progenitors ◽  
Histamine Synthesis ◽  
Histamine Production ◽  
Dependent Cell

The calcium ionophore A23187 promotes histamine synthesis in murine bone marrow cells by increasing the expression of mRNA encoding histidine decarboxylase (HDC), the histamine-forming enzyme. The cells responsible for this biological activity copurify with hematopoietic progenitors in terms of density, light scatter characteristics, and rhodamine retention, similar to interleukin (IL) 3-induced histamine- producing cells. Yet, the effect of calcium ionophore is not mediated by IL-3. The most purified rhodamine-bright bone marrow subset contains 80% cells that respond to calcium ionophore by increased HDC mRNA expression. This high frequency makes the involvement of one particular progenitor subset in histamine synthesis unlikely. The finding that all IL-3-dependent cell lines tested so far exhibit increased histamine production and HDC mRNA expression in response to calcium influx lends further support to this notion. Cell lines requiring other growth factors or proliferating spontaneously lack this ability. Finally, it should be noted that IL-3-dependent cell lines do not produce histamine in response to their growth factor. It might, therefore, be suggested that the pathway transducing the signal for increased histamine synthesis after IL-3 receptor binding in normal hematopoietic progenitors is modified in these cell lines.

scholarly journals Platelet Factor 4 and Other CXC Chemokines Support the Survival of Normal Hematopoietic Cells and Reduce the Chemosensitivity of Cells to Cytotoxic Agents

Blood ◽  
1997 ◽  
Vol 89 (7) ◽  
pp. 2328-2335 ◽  
Author(s):  
Zhong Chao Han ◽  
Min Lu ◽  
Junmin Li ◽  
Mai Defard ◽  
Bernadette Boval ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Growth Factor ◽  
Cell Lines ◽  
Cancer Cell ◽  
Bone Marrow Cells ◽  
Hematopoietic Cells ◽  
Platelet Factor 4 ◽  
Cytotoxic Agents ◽  
Platelet Factor ◽  
Cd34 Cells

Abstract The effects of platelet factor 4 (PF4) on the viability and chemosensitivity of normal hematopoietic cells and cancer cell lines were studied to determine the mechanisms whereby PF4 functions as either an inhibitor or a protector and to evaluate its clinical significance. Two other chemokines, interleukin-8 (IL-8) and neutrophil-activating peptide-2 (NAP-2), were also studied in comparison to PF4. Using a tetrazolium salt assay for cell viability, we observed that PF4 at 1 to 50 μg/mL supported the viability of normal human bone marrow cells. Approximately 45% of cells cultured for 48 hours survived, whereas 80% or more survived in the presence of PF4 5 μg/mL. PF4 also supported the viability of CD34+ cord blood (CB) cells and protected them from apoptosis induced by transforming growth factor β1 (TGFβ1) and cytotoxic drugs. Pretreatment of CD34+ cells by PF4, but not by TGFβ1, caused an increase in the number of megakaryocyte colonies after these cells were replated in secondary cultures. Flow cytometry analysis showed that when CD34+ cells were preincubated with PF4 or TGFβ1 for 12 days in hematopoietic growth factor–rich medium, an increased number of remaining CD34+ cells was observed only for PF4-treated cells. Furthermore, PF4 significantly reduced the chemosensitivity of bone marrow cells, as shown by its ability to increase the 50% inhibition concentration (IC50) of several cytotoxic agents. Like PF4, IL-8 and NAP-2 at 0.1, 0.6, and 1 μg/mL supported the survival of myeloid progenitors, including colony-forming units granulocyte, erythroblast, monocyte, megakaryocyte (CFU-GEMM), CFU-megakaryocyte (CFU-MK), CFU–granulocyte/macrophage (CFU-GM), and burst-forming units–erythroblast (BFU-E), and reduced their sensitivity to the toxicity of etoposide (ETP). Protamine sulfate at 1 to 100 μg/mL showed no such activity of PF4. Interestingly, the three chemokines failed to affect significantly the viability and chemosensitivity of three leukemic and two other tumor cell lines. Based on these results, we conclude for the first time that PF4 and IL-8 and NAP-2 support the survival of normal hematopoietic precursors and protect them from the toxicity of chemotherapeutic agents. Because such activities are unique to normal hematopoietic cells but not to the cancer cell lines evaluated, a potential clinical application of these molecules in the treatment of cancer is suggested.

PTEN Expression Is Decreased in Pediatric Burkitt Lymphoma Tissues but Does Not Correlate with AKT Activation.

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 2664-2664
Author(s):  
Rodic Vladimir ◽  
Tripp Sheryl ◽  
Michael G Bayerl ◽  
Phillip Barnette ◽  
Joshua D. Schiffman ◽  
...  
Keyword(s):  
Lymph Node ◽  
Mrna Expression ◽  
Cell Lines ◽  
Burkitt Lymphoma ◽  
Pten Expression ◽  
Mrna Levels ◽  
Pten Protein ◽  
Akt Activation ◽  
Pten Mrna ◽  
Reactive Lymph Node

Abstract Abstract 2664 Background: We have recently shown that a subset of pediatric Burkitt lymphoma (pBL) show gain of 13q31, containing the microRNA (miRNA) MIR-17–92 locus, as well as associated increased expression of miRNA from that locus. PTEN, a tumor suppressor phosphatase that negatively regulates the PI3K/AKT/mTOR pro-survival pathway, is negatively regulated by MIR-17–92. Therefore, we hypothesized that PTEN protein and mRNA expression would be decreased in pBL cases with 13q31 locus gain and high expression of MIR-17–92. Because PTEN negatively regulates the PI3K/AKT/mTOR pathway, we also assessed AKT activation in pBL tissues. Methods: We assessed PTEN protein expression by immunohistochemistry (IHC) in formalin-fixed, paraffin-embedded (FFPE) pBL tissues from a cohort of patients (n=25) where MIR-17–92 expression levels and status of the 13q31 locus have been previously determined (Schiffman/Miles, Brit J Haematol 155:477, 2011). From FFPE tissues, we assayed PTEN mRNA levels by quantitative RT-PCR. The results of protein and mRNA expression were then correlated with presence or absence of a 13q31 gain and the expression level of MIR17. To evaluate AKT activation in pBL, we assayed for phospho-(p)-AKT by IHC. In addition, PTEN, AKT, and p-AKT protein levels were evaluated by western blotting in Raji and Ramos BL cell lines. Results: PTEN protein was weakly expressed in 5/25 cases (20%) of pBL. PTEN protein showed moderate nuclear and cytoplasmic staining in a subset of paracortical and germinal center lymphocytes in reactive lymph node control tissues, and vascular endothelial cells provided internal positive control in tumor tissue. pBL tissues showed decreased relative PTEN mRNA expression compared to reactive lymph node controls (1.56 vs. 2.61, p<3E-07). There was no significant difference in PTEN mRNA levels between PTEN protein positive and negative cases (1.27 vs. 1.61, p=0.22); cases with or without 13q31gain (1.49 vs. 1.57, p=0.60); or cases with high or low expression of MIR17. p-AKT was detected by IHC in 1/25 cases (4%). The BL cell lines Raji and Ramos showed very low levels of PTEN protein but no detectable p-AKT; total AKT was readily detectable. Discussion: pBL shows decreased PTEN mRNA, and most cases are negative for PTEN protein expression. Unexpectedly, higher MIR-17–92 expression level or 13q31 gain did not show an inverse relationship with PTEN expression at the protein or mRNA level. Despite decreased PTEN expression, p-AKT was detected in only 1/25 pBL cases and was not detected in BL cell lines. These findings suggest that decreased PTEN expression does not lead to AKT activation in pBL. Ongoing studies will investigate the potential of alternate PTEN targets and/or alternate targets for MIR-17–92 in pBL. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Growth factors increase amphotropic retrovirus binding to human CD34+ bone marrow progenitor cells

Blood ◽  
1993 ◽  
Vol 82 (11) ◽  
pp. 3290-3297 ◽  
Author(s):  
GM Crooks ◽  
DB Kohn
Keyword(s):  
Bone Marrow ◽  
Growth Factors ◽  
Gene Transfer ◽  
Hematopoietic Cells ◽  
Receptor Expression ◽  
Cell Surface Receptor ◽  
Virus Binding ◽  
Steel Factor ◽  
Human Cd34 ◽  
Cd34 Cells

Gene transfer into human cells using murine amphotropic retroviral vectors is the basic technique used in most current gene therapy studies. The identity of the cell surface receptor for the amphotropic envelope remains unknown and thus its importance in gene transfer is poorly understood. We have measured specific retrovirus binding to cells to study amphotropic virus receptor regulation in human CD34+ bone marrow (BM) progenitors and primitive CD34+CD38- human hematopoietic cells. The rat monoclonal antibody 83A25 recognizes an epitope common to the envelope glycoprotein of all classes of Moloney murine leukemia virus. Indirect fluorescent labeling of 83A25 allows flow cytometric analysis of specific virus-cell interactions and is an indirect measure of specific receptors. Using this assay, amphotropic virus binding to fresh CD34+ cells was minimal. However, when CD34+ cells were cultured with or without growth factors for 4 days, specific binding of amphotropic retrovirus was readily shown. Inclusion of interleukin-3 (IL-3), IL-6, and Steel factor in cultures increased the fluorescence associated with amphotropic virus binding by twofold to four-fold (mean fold increase 2.7 +/- 0.84). Virus binding to CD34+CD38- cells was shown only in those cells culture in IL-3, IL-6, and Steel factor. These results suggest that certain cytokines may cause an increase in the number and/or affinity of amphotropic receptors on primitive human hematopoietic cells. Upregulation of viral receptor expression may be one of the mechanisms by which cytokines enhance gene transfer into primitive BM cells.

scholarly journals CHARACTERIZATION OF COLLAGEN (IV) MRNA IN CELL LINES OF BREAST CANCER

2015 ◽  
Vol 77 (25) ◽  
Author(s):  
Afzan Mat Yusof ◽  
Mardhiah Mohammad ◽  
Sharifah Norbaizura Syed Bahrom ◽  
Syahirah Kaja Mohideen ◽  
Ridhwan Roshdi ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Mrna Expression ◽  
Cancer Cells ◽  
Cell Lines ◽  
Cancer Cell ◽  
Breast Cancer Cell ◽  
Cancer Cell Lines ◽  
Collagen Iv ◽  
Breast Cancer Cell Lines ◽  
Mrna Levels

Breast cancer incidence rate has increased in the 5 recent years with 14% increases in mortality. The structural change in the collagen chain has led to alterations in the cancer cells. Various biological processes, such as differentiation or gene expression, are regulated through extracelullar matrix (ECM)[1]. The restructuring of the collagenous architecture in the hypoxic microenvironment may influence the invasive growth of the cancer cells. With the increased stress within the cell, the invasion of cancer cells into the ECM was triggered. This cell lines model would enable the exploration of the relationship between the extracellular matrices component and the tumor proliferation. The aim of this study is to characterize the collagen (IV) mRNA expression in the breast cancer cell.  Breast cancer (MCF7) cell lines were cultured and harvested upon confluent. The RNA was extracted from the cell lines and then the cDNA were synthesized. The collagen (IV) mRNA levels in breast cancer cell lines were measured using real time PCR and GAPDH was used as an internal control. The level of COL4A2 (IV) mRNA expression was higher compared with COL4A1 (IV) mRNA. The level of COL4A5 (IV) mRNA was reduced significantly in breast cancer cells lines. Overall, the expression of COL4A1-A6 (IV) was reduced. The reduced amount of collagen (IV) in breast cancer cell lines suggested that the collagen was restructured and this has triggered the tumor invasion into the ECM.

Expression of Macrophage Inflammatory Protein-1 Receptors in Human CD34+ Hematopoietic Cells and Their Modulation by Tumor Necrosis Factor- and Interferon-γ

Blood ◽  
1998 ◽  
Vol 92 (9) ◽  
pp. 3073-3081 ◽  
Author(s):  
Jan Dürig ◽  
Erika A. de Wynter ◽  
Christoph Kasper ◽  
Michael A. Cross ◽  
James Chang ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Tumor Necrosis ◽  
Hematopoietic Cells ◽  
Interferon Γ ◽  
Receptor Expression ◽  
Receptor Subtypes ◽  
Inflammatory Protein ◽  
Cd34 Cells ◽  
Macrophage Inflammatory Protein ◽  
Necrosis Factor

Abstract Macrophage inflammatory protein-1 (MIP-1) can stimulate growth inhibitory and potent chemotactic functions in hematopoietic cells. To investigate whether the action of MIP-1 may be regulated at the cellular receptor level, we studied the expression and modulation of MIP-1 receptors on CD34+ cells isolated from normal bone marrow (NBM), umbilical cord blood (CB), and leukapheresis products (LP). Expression of MIP-1 receptors on CD34+cells was analyzed by two-color flow cytometry using a biotinylated MIP-1 molecule. The mean percentage of LP CD34+ cells expressing the MIP-1 receptors was 67.7 ± 7.2% (mean ± SEM; n = 22) as compared with 89.9 ± 2.6% (n = 10) and 74.69 ± 7.04% (n = 10) in CB and NBM, respectively (P = .4). The expression of the MIP-1 receptor subtypes on LP CD34+ cells was studied by indirect immunofluorescence using specific antibodies for the detection of CCR-1, CCR-4, and CCR-5. Microscopical examination revealed a characteristic staining of the cytoplasmic cell membrane for all three receptor subtypes. Detailed analysis of two LP samples showed that 65.8%, 4.4%, and 30.5% of CD34+ cells express CCR-1, CCR-4, and CCR-5, respectively. Culture of LP CD34+ cells for 24 to 36 hours in the presence of tumor necrosis factor- (TNF-) and interferon-γ (IFN-γ) resulted in a significant increase in MIP-1 receptor expression. TNF- induced MIP-1 receptor upregulation in a time- and concentration-dependent manner. Our results suggest that inhibitory cytokines produced by the bone marrow microenvironment are likely to be involved in the regulation of MIP-1 receptor expression on hematopoietic cells. © 1998 by The American Society of Hematology.

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