scholarly journals Regulation of the interleukin-3 (IL-3) receptor by IL-3 in the fetal liver-derived FL5.12 cell line

Blood ◽  
1994 ◽  
Vol 83 (9) ◽  
pp. 2459-2468 ◽  
Author(s):  
PA Algate ◽  
LS Steelman ◽  
MW Mayo ◽  
A Miyajima ◽  
JA McCubrey
Keyword(s):  
Cell Lines ◽  
Fetal Liver ◽  
Receptor Expression ◽  
Mrna Levels ◽  
Interleukin 3 ◽  
Rna Transcripts ◽  
Rna Analysis ◽  
Cognate Receptor ◽  
Parental Factor ◽  
Low Levels

To determine the effects of a cytokine on cognate receptor expression in normal and neoplastic cells, the interleukin-3 receptor (IL-3R) complex was examined in the parental IL-3-dependent line FL5.12, which was isolated from fetal liver, and in autocrine- and v-abl-transformed derivative lines. IL-3 decreased the amount of the IL-3R alpha and beta chains detected on the cell surface of the parental IL-3-dependent cells. In contrast, high levels of IL-3R beta were constitutively detected on the autocrine-transformed lines in the absence and presence of exogenous IL-3. Only low levels of IL-3R beta were observed in the two v-abl-transformed derivative cell lines examined, which no longer required IL-3 for growth. The levels of the IL-3R alpha chain detected were similar in these transformed cells and were not regulated by IL-3. These results were substantiated further by RNA analysis, because IL-3 decreased the levels of IL-3R transcripts in the parental factor- dependent FL5.12 line. The pattern of IL-3R gene expression was opposite to that of other receptors or proto-oncogenes, because RNA transcripts for all other genes examined were induced by IL-3. We conclude that IL-3 tightly controls IL-3R expression in the IL-3- dependent FL5.12 cells, whereas steady-state mRNA levels were not altered in the two v-abl-transformed derivative cell lines examined in this study.

scholarly journals Regulation of the interleukin-3 (IL-3) receptor by IL-3 in the fetal liver-derived FL5.12 cell line

Blood ◽  
1994 ◽  
Vol 83 (9) ◽  
pp. 2459-2468 ◽  
Author(s):  
PA Algate ◽  
LS Steelman ◽  
MW Mayo ◽  
A Miyajima ◽  
JA McCubrey
Keyword(s):  
Cell Lines ◽  
Fetal Liver ◽  
Receptor Expression ◽  
Mrna Levels ◽  
Interleukin 3 ◽  
Rna Transcripts ◽  
Rna Analysis ◽  
Cognate Receptor ◽  
Parental Factor ◽  
Low Levels

Abstract To determine the effects of a cytokine on cognate receptor expression in normal and neoplastic cells, the interleukin-3 receptor (IL-3R) complex was examined in the parental IL-3-dependent line FL5.12, which was isolated from fetal liver, and in autocrine- and v-abl-transformed derivative lines. IL-3 decreased the amount of the IL-3R alpha and beta chains detected on the cell surface of the parental IL-3-dependent cells. In contrast, high levels of IL-3R beta were constitutively detected on the autocrine-transformed lines in the absence and presence of exogenous IL-3. Only low levels of IL-3R beta were observed in the two v-abl-transformed derivative cell lines examined, which no longer required IL-3 for growth. The levels of the IL-3R alpha chain detected were similar in these transformed cells and were not regulated by IL-3. These results were substantiated further by RNA analysis, because IL-3 decreased the levels of IL-3R transcripts in the parental factor- dependent FL5.12 line. The pattern of IL-3R gene expression was opposite to that of other receptors or proto-oncogenes, because RNA transcripts for all other genes examined were induced by IL-3. We conclude that IL-3 tightly controls IL-3R expression in the IL-3- dependent FL5.12 cells, whereas steady-state mRNA levels were not altered in the two v-abl-transformed derivative cell lines examined in this study.

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 Interleukin-3, GM-CSF, and G-CSF receptor expression on cell lines and primary leukemia cells: receptor heterogeneity and relationship to growth factor responsiveness

Blood ◽  
1989 ◽  
Vol 74 (1) ◽  
pp. 56-65 ◽  
Author(s):  
LS Park ◽  
PE Waldron ◽  
D Friend ◽  
HM Sassenfeld ◽  
V Price ◽  
...  
Keyword(s):  
Growth Factor ◽  
Cell Lines ◽  
Specific Binding ◽  
Absolute Number ◽  
Lymphocytic Leukemia ◽  
Receptor Expression ◽  
Interleukin 3 ◽  
Gm Csf ◽  
In Vitro Cell Lines

Abstract Recombinant human granulocyte-macrophage (GM) colony-stimulating factor (GM-CSF), G-CSF, and interleukin-3 (IL-3) labeled with 125I were used to study the characteristics and distribution of receptors for these factors on in vitro cell lines and on cells from patients with acute nonlymphocytic leukemia (ANL) and acute lymphocytic leukemia (ALL). Receptors for GM-CSF and G-CSF were restricted to a subset of myelomonocytic cell lines whereas IL-3 receptors were also found on pre- B- or early B-cell lines. Receptors for all three CSFs were broadly distributed on ANL cells, with considerable variability in levels of expression. Measurement of the colony-forming ability of ANL cells in response to the CSFs showed that there was no direct correlation between the ability of the cells to respond to a growth factor and the absolute number of receptors expressed for that growth factor. Binding of radiolabeled IL-3 and GM-CSF to ANL cells produced complex biphasic curves. Further analysis showed that both IL-3 and GM-CSF were able to partially compete for specific binding of the heterologous radiolabeled ligand to cells from several ANL patients, suggesting that heterogeneity may exist in human CSF receptors. These results provide new insights into the complex role that CSFs may play in ANL.

scholarly journals CSIG-18. NERVE/GLIAL ANTIGEN (NG)2 EXPRESSION IN GLIOBLASTOMA IS REGULATED BY miR-29b-3p

2019 ◽  
Vol 21 (Supplement_6) ◽  
pp. vi47-vi48
Author(s):  
Beate Schmitt ◽  
Anne Boewe ◽  
Yuan Gu ◽  
Christoph Sippl ◽  
Steffi Urbschat ◽  
...  
Keyword(s):  
Drug Resistance ◽  
Cell Lines ◽  
Therapeutic Strategy ◽  
Mrna Levels ◽  
Glioblastoma Cell ◽  
Glioblastoma Cells ◽  
Qrt Pcr ◽  
Glioblastoma Patient ◽  
Low Levels ◽  
Glioblastoma Cell Lines

Abstract Overexpression of NG2 in human glioblastoma cells is associated with an elevated drug resistance and thereby worsens clinical outcome. However, the regulatory mechanisms of NG2 expression are largely unknown. In this study, we identified miR-29b-3p as a posttranscriptional factor of NG2 expression. The basal mRNA levels of miR-29b-3p and NG2 were detected in the NG2-positive glioblastoma cell lines A1207 and U87 by qRT-PCR. The cells were transfected with miR-29b-3p-mimic or scrambled-miR (control) and the expression of NG2 was analyzed by qRT-PCR, flow cytometry and Western blot. Reporter gene analyses of the NG2 promotor region and 3’UTR were performed to study the effect of miR-29b-3p on NG2 expression. Finally, we analyzed the mRNA levels of NG2 and miR-29b-3p in samples from glioblastoma patients. We found that the two NG2-positive glioblastoma cell lines A1207 and U87 are positive for miR-29b-3p. Transfection with miR-29b-3p-mimic reduced NG2 mRNA levels in A1207 (29%±9.9; Mean±SD) and U87 (6%±2.8), resulting in a significantly decreased NG2 protein expression in A1207 (67%±6.4) and U87 (75%±4) when compared to controls. The analysis of the 3’UTR revealed that miR-29b-3p is a posttranscriptional regulator of NG2 expression. Moreover, miR-29b-3p affects the pretranscriptional NG2 expression by diminishing SP-1-dependent NG2 promotor activity. These results were confirmed by the analysis of glioblastoma patient-derived samples, demonstrating that a high NG2 expression is associated with low levels of miR-29b-3p. In conclusion, we identified miR-29b-3p as a crucial regulator of NG2 expression in glioblastoma. Hence, targeting NG2 expression by miR-29b-3p may provide a novel therapeutic strategy to overcome drug resistance in NG2-positive glioblastoma cells.

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.

scholarly journals Interleukin-3, GM-CSF, and G-CSF receptor expression on cell lines and primary leukemia cells: receptor heterogeneity and relationship to growth factor responsiveness

Blood ◽  
1989 ◽  
Vol 74 (1) ◽  
pp. 56-65 ◽  
Author(s):  
LS Park ◽  
PE Waldron ◽  
D Friend ◽  
HM Sassenfeld ◽  
V Price ◽  
...  
Keyword(s):  
Growth Factor ◽  
Cell Lines ◽  
Specific Binding ◽  
Absolute Number ◽  
Lymphocytic Leukemia ◽  
Receptor Expression ◽  
Interleukin 3 ◽  
Gm Csf ◽  
In Vitro Cell Lines

Recombinant human granulocyte-macrophage (GM) colony-stimulating factor (GM-CSF), G-CSF, and interleukin-3 (IL-3) labeled with 125I were used to study the characteristics and distribution of receptors for these factors on in vitro cell lines and on cells from patients with acute nonlymphocytic leukemia (ANL) and acute lymphocytic leukemia (ALL). Receptors for GM-CSF and G-CSF were restricted to a subset of myelomonocytic cell lines whereas IL-3 receptors were also found on pre- B- or early B-cell lines. Receptors for all three CSFs were broadly distributed on ANL cells, with considerable variability in levels of expression. Measurement of the colony-forming ability of ANL cells in response to the CSFs showed that there was no direct correlation between the ability of the cells to respond to a growth factor and the absolute number of receptors expressed for that growth factor. Binding of radiolabeled IL-3 and GM-CSF to ANL cells produced complex biphasic curves. Further analysis showed that both IL-3 and GM-CSF were able to partially compete for specific binding of the heterologous radiolabeled ligand to cells from several ANL patients, suggesting that heterogeneity may exist in human CSF receptors. These results provide new insights into the complex role that CSFs may play in ANL.

Eltrombopag Decreases Proliferation of Ovarian, Lung and Breast Tumor Cell Lines.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 2409-2409
Author(s):  
Connie L. Erickson-Miller ◽  
Jennifer Kirchner ◽  
Kodandaram Pillarisetti ◽  
Lone Ottesen ◽  
Yasser Mostafa Kamel ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Tumor Cell ◽  
Cell Lines ◽  
Mrna Levels ◽  
Tumor Cell Lines ◽  
Employment Equity ◽  
Qrt Pcr ◽  
Thrombopoietin Receptor ◽  
Equity Ownership ◽  
Low Levels

Abstract Abstract 2409 Poster Board II-386 Background: Eltrombopag (Promacta®) is a novel, oral thrombopoietin receptor (TpoR) agonist that interacts with the TpoR on bone marrow progenitors to stimulate megakaryocyte production, thus increasing platelet counts in thrombocytopenic patients. The effects of eltrombopag on the proliferation of solid tumor cell lines and the expression of thrombopoietin receptor (MPL, TpoR) on patient tumors is of interest given that chemotherapy can cause thrombocytopenia. Materials and methods: Proliferation was measured by Cell Titer Glo assay on 3 ovarian (OVCAR3, OVCAR4, SKOV3), 4 lung (A549, NCI-H226, NCI-H510, NCI-H460) and 3 breast (BT-474, MCF7, HCC1937) cancer cell lines from the ATCC treated with 0.01 – 100 ug/mL eltrombopag. Quantitative RT-PCR (qRT-PCR) for MPL expression was performed on the tumor cell lines and on 40 tumor samples, each from subjects with ovarian, lung or breast cancer. Microarray analysis for MPL mRNA expression was examined from 118 subjects with breast cancer and 29 with non-small cell lung cancer (NSCLC). Microarray data was normalized using robust multiarray average (RMA) and relative mRNA expression was determined. To determine expression of TpoR protein, western blot analyses was performed on some of the tumor cell lines. Results: Eltrombopag induced an inhibition of proliferation on all of the ovarian, lung and breast solid tumor cell lines tested. The IC50 ranged from 3.7 to 49.7 ug/mL (see table below). The Cmax of ITP patients treated with 75 mg eltrombopag is 11.4 ug/mL, demonstrating that these concentrations are clinically achievable. There was no enhancement of proliferation at any concentration of eltrombopag, consistent with the very low or undetectable level of MPL expression on samples of tumors from patients with these diseases. MPL was expressed at very low or undetectable levels in these tumor cell lines with the exception of the lung cancer line, NCI-H510. However, western blot analyses showed no detectable TpoR protein expression regardless of the higher levels of MPL mRNA in NCI-H510 cells. Erythropoietin receptor (EPOR) mRNA was expressed at low-to-moderate levels, while ERBB2 and IGF1R were expressed at higher levels in these cell lines. Microarray analysis showed undetectable MPL mRNA levels in all 118 samples from patients with breast cancer and 52% of the NSCLC samples, the remaining NSCLC samples expressed low levels of MPL. In contrast, EPOR was expressed in 75–100% of the breast cancer, and NSCLC samples. ERBB2 was expressed in 97–100% of the samples and IGF1R was expressed in 54–100% of the samples. When 40 other tumor samples each from subjects with ovarian, lung and breast cancer were examined by qRT-PCR, MPL mRNA levels were also very low or undetectable. EPOR, ERBB2, and IGF1R expression levels varied according to tumor type, but were greater than MPL levels. Conclusions: In summary, similar to its effects on leukemia and lymphoma cell lines, all of the nine lung, ovarian, breast or prostate tumor cell lines demonstrated decreased proliferation in response to eltrombopag. The undetectable or very low levels of expression of MPL mRNA in tumors of patients with lung, ovarian, breast or prostate cancer supports the proliferation results. Disclosures: Erickson-Miller: GlaxoSmithKline: Employment, Equity Ownership, Patents & Royalties, Research Funding. Kirchner:GlaxoSmithKline: Employment. Pillarisetti:GSK: Employment, Equity Ownership, Patents & Royalties. Ottesen:GSK: Employment, Equity Ownership. Mostafa Kamel:GSK: Employment, Equity Ownership. Liu:GSK: Employment, Equity Ownership. Martin:GSK: Employment, Equity Ownership. Messam:GSK: Employment, Equity Ownership.

scholarly journals RELA is Sufficient to Mediate Interleukin-1 (IL-1) Repression of Androgen Receptor Expression and Activity in LNCaP Disease Progression Model

2019 ◽  
Author(s):  
Shayna E. Thomas-Jardin ◽  
Haley Dahl ◽  
Mohammed S. Kanchwala ◽  
Freedom Ha ◽  
Joan Jacob ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Androgen Receptor ◽  
Disease Progression ◽  
Cell Lines ◽  
Pathway Analysis ◽  
Molecular Mechanisms ◽  
Interleukin 1 ◽  
Receptor Expression ◽  
Mrna Levels ◽  
Lncap Cells

ABSTRACTBackgroundThe Androgen Receptor (AR) nuclear transcription factor is a therapeutic target for prostate cancer (PCa). Unfortunately, patients can develop resistance to AR-targeted therapies and progress to lethal disease, underscoring the importance of understanding the molecular mechanisms that underlie treatment resistance. Inflammation is implicated in PCa initiation and progression and we have previously reported that the inflammatory cytokine, interleukin-1 (IL-1), represses AR mRNA levels and activity in AR-positive (AR+) PCa cell lines concomitant with the upregulation of pro-survival biomolecules. Thus, we contend that IL-1 can select for AR-independent, treatment-resistant PCa cells.MethodsTo begin to explore how IL-1 signaling leads to the repression of AR mRNA levels, we performed comprehensive pathway analysis on our RNA sequencing data from IL-1-treated LNCaP PCa cells. Our pathway analysis predicted Nuclear Factor Kappa B (NF-κB) p65 subunit (RELA), a canonical IL-1 signal transducer, to be significantly active and potentially regulate many genes, including AR. We used siRNA to silence the NF-κB family of transcription factor subunits, RELA, RELB, c-REL, NFKB1, or NFKB2, in IL-1-treated LNCaP, C4-2, and C4-2B PCa cell lines. C4-2 and C4-2B cell lines are castration-resistant LNCaP sublines and represent progression towards metastatic PCa disease; and we have previously shown that IL-1 represses AR mRNA levels in C4-2 and C4-2B cells.ResultssiRNA revealed that RELA alone is sufficient to mediate IL-1 repression of AR mRNA and AR activity. Intriguingly, while LNCaP cells are more sensitive to IL-1-mediated repression of AR than C4-2 and C4-2B cells, RELA siRNA led to a more striking de-repression of AR mRNA levels and AR activity in C4-2 and C4-2B cells than in LNCaP cells.ConclusionsThese data indicate that there are RELA-independent mechanisms that regulate IL-1-mediated AR repression in LNCaP cells and suggest that the switch to RELA-dependent IL-1 repression of AR in C4-2 and C4-2B cells reflects changes in epigenetic and transcriptional programs that evolve during PCa disease progression.

scholarly journals Prognostic value of TERF1 expression in prostate cancer

2021 ◽  
Vol 33 (1) ◽  
Author(s):  
Gabriel Arantes dos Santos ◽  
Nayara Izabel Viana ◽  
Ruan Pimenta ◽  
Vanessa Ribeiro Guimarães ◽  
Juliana Alves de Camargo ◽  
...  
Keyword(s):  
Prostate Cancer ◽  
Overall Survival ◽  
Cell Lines ◽  
Normal Tissue ◽  
Receptor Expression ◽  
Telomere Maintenance ◽  
Localized Prostate Cancer ◽  
Cancer Tissue ◽  
Mrna Levels ◽  
Clinical Specimens

Abstract Background Telomere dysfunction is one of the hallmarks of cancer and is crucial to prostate carcinogenesis. TERF1 is a gene essential to telomere maintenance, and its dysfunction has already been associates with several cancers. TERF1 is a target of miR-155, and this microRNA can inhibit its expression and promotes carcinogenesis in breast cancer. We aim to analyze TERF1, in gene and mRNA level, involvement in prostate cancer progression. Results Alterations in TERF1 DNA were evaluated using datasets of primary tumor and castration-resistant tumors (CRPC) deposited in cBioportal. The expression of TERF1 mRNA levels was assessed utilizing TCGA datasets, clinical specimens, and metastatic prostate cancer cell lines (LNCaP, DU145, and PC3). Six percent of localized prostate cancer presents alterations in TERF1 (the majority of that was amplifications). In the CRPC cohort, 26% of samples had TERF1 amplification. Patients with TERF1 alterations had the worst overall survival only on localized cancer cohort (p = 0.0027). In the TCGA cohort, mRNA levels of TERF1 were downregulated in comparison with normal tissue (p = 0.0013) and upregulated in tumors that invade lymph nodes (p = 0.0059). The upregulation of TERF1 is also associated with worst overall survival (p = 0.0028) and disease-free survival (p = 0.0023). There is a positive correlation between TERF1 and androgen receptor expression in cancer tissue (r = 0.53, p < 0.00001) but not on normal tissue (r = − 0.16, p = 0.12). In the clinical specimens, there is no detectable expression of TERF1 and upregulation of miR-155 (p = 0.0348). In cell lines, TERF1 expression was higher in LNCaP and was progressively lower in DU145 and PC3 (p = 0.0327) with no differences in miR-155 expression. Conclusion Amplification/upregulation of TERF1 was associated with the worst prognostic in localized prostate cancer. Our results corroborate that miR-155 regulates TERF1 expression in prostate cancer. TERF1 has the potential to become a biomarker in prostate cancer.

Molecular and functional analysis of glutamine uptake in human hepatoma and liver-derived cells

2002 ◽  
Vol 283 (5) ◽  
pp. G1062-G1073 ◽  
Author(s):  
Barrie P. Bode ◽  
Bryan C. Fuchs ◽  
Bryan P. Hurley ◽  
Jennifer L. Conroy ◽  
Julie E. Suetterlin ◽  
...  
Keyword(s):  
Amino Acid ◽  
Cell Lines ◽  
Competitive Inhibition ◽  
Fetal Liver ◽  
Hepatoma Cell ◽  
Human Hepatocytes ◽  
Epithelial Cell Line ◽  
Mrna Levels ◽  
Human Hepatoma ◽  
Glutamine Uptake

Human hepatoma cells take up glutamine at rates severalfold faster than the system N-mediated transport rates observed in normal human hepatocytes. Amino acid inhibition, kinetic, Northern blotting, RT-PCR, and restriction enzyme analyses collectively identified the transporter responsible in six human hepatoma cell lines as amino acid transporter B0(ATB0), the human ortholog of rodent ASCT2. The majority of glutamine uptake in liver fibroblasts and an immortalized human liver epithelial cell line (THLE-5B) was also mediated by ATB0. The 2.9-kb ATB0 mRNA was equally expressed in all cell lines, whereas expression of the system A transporters ATA2 and ATA3 was variable. In contrast, the system N isoforms (SN1 and SN2) were expressed only in well-differentiated hepatomas. ATB0 mRNA was also expressed in cirrhotic liver and adult and pediatric liver cancer biopsies but was not detectable in isolated human hepatocytes or fetal liver. Although the growth of all hepatomas was glutamine dependent, competitive inhibition of ATB0-mediated glutamine uptake blocked proliferation only in poorly differentiated cells lacking SN1 or SN2 expression and exhibiting low glutamine synthetase mRNA levels.

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