Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Differentiated vs. undifferentiated promyelocytic cell lines (HL-60 and KG-1) and Neutrophiles

ABSTRACT: We identify the tetraspanin CD82 as the recognition site for xenogeneic endothelial cells independently of Gala1,3-gal structures. We demonstrate that in contrast to undifferentiated cells, differentiated promyelocytic cell lines (HL-60, THP-1 and KG-1) are capable of recognizing xenogeneic porcine endothelial cells in a calcium-dependent manner. We used serial analysis of gene expression (SAGE) to identify the differentially expressed transcripts in these cell lines. Interrogation of these transcripts revealed a number of differentially expressed genes that include the Gala1,3-gal-independent recognition moiety(s). Comparing these SAGE transcripts with those expressed in resting human naive neutrophils identified the tetraspanin CD82 as the most likely candidate responsible for xenogeneic recognition. Blocking antibodies to CD82 in human naive neutrophils inhibited the calcium response and abolished the subsequent Reactive Oxygen Metabolite (ROM) production evoked by the xenogeneic encounter of either Gala1,3-gal knockout or wild-type porcine aortic endothelial cells. Our data identify CD82 on innate immune cells as the major recognition moiety of the xenogeneic endothelium, independently of Gala1,3-gal-structures and open new avenues of intervention to making xenotransplantation a clinical reality. We used serial analysis of gene expression (SAGE) to identify the differentially expressed transcripts in promyelocytic cell lines (HL-60, THP-1 and KG-1) upon differentiation. We looked for genes common to the differentiation pathway in these cell lines but absent from the control naiive neutrophiles.

ORGANISM(S): Homo sapiens

SUBMITTER: Anason Halees

PROVIDER: E-GEOD-43211 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

Identification of the tetraspanin CD82 as a new barrier to xenotransplantation.

Saleh Soad M SM Parhar Ranjit S RS Al-Hejailan Reem S RS Bakheet Razan H RH Khaleel Hala S HS Khalak Hanif G HG Halees Anason S AS Zaidi Marya Z MZ Meyer Brian F BF Yung Gisella P GP Seebach Jörg D JD Conca Walter W Khabar Khalid S KS Collison Kate S KS Al-Mohanna Futwan A FA

Journal of immunology (Baltimore, Md. : 1950) 20130719 5

Significant immunological obstacles are to be negotiated before xenotransplantation becomes a clinical reality. An initial rejection of transplanted vascularized xenograft is attributed to Galα1,3Galβ1,4GlcNAc-R (Galα1,3-Gal)-dependent and -independent mechanisms. Hitherto, no receptor molecule has been identified that could account for Galα1,3-Gal-independent rejection. In this study, we identify the tetraspanin CD82 as a receptor molecule for the Galα1,3-Gal-independent mechanism. We demonstra ...[more]

PMID: 23872050

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Project description:We recently found that the tetraspanin family member, CD82, which is aberrantly expressed in chemotherapy-resistant CD34+/CD38− acute myelogenous leukemia (AML) cells, negatively regulates matrix metalloproteinase 9, and plays an important role in enabling CD34+/CD38− AML cells to adhere to the bone marrow microenvironment. This study explored novel functions of CD82 that contribute to AML progression. We employed microarray analysis comparing the gene expression profiles between CD34+/CD38− AML cells transduced with CD82 shRNA and CD34+/CD38− AML cells transduced with control shRNA. Real-time RT-PCR and western blot analysis were performed to examine the effect of CD82 knockdown on the expression of the polycomb group member, enhancer of zeste homolog 2 (EZH2), in leukemia cells. A chromatin immunoprecipitation assay was performed to examine the effect of CD82 expression on the amount of EZH2 bound to the promoter regions of tumor suppressor genes in leukemia cells. We also utilized methylation-specific PCR to examine whether CD82 expression influences the methylation status of the tumor suppressor gene promoter regions in leukemia cells. Microarray analysis revealed that levels of EZH2 decreased after shRNA-mediated depletion of CD82 in CD34+/CD38− AML cells. Moreover, the antibody-mediated blockade of CD82 in leukemia cells lowered EZH2 expression via activation of p38 MAPK signaling, decreased the amount of EZH2 bound to the promoter regions of the tumor suppressor genes, and inhibited histone H3 lysine 27 trimethylation in these promoter regions, resulting in upregulation of the tumor suppressors at both the mRNA and protein levels. The aim of this study was to explore the biological functions of a tetraspanin family protein––CD82–– expressed aberrantly in chemotherapy-resistant CD34+/CD38- acute myelogenous leukemia (AML) cells. Microarray analysis of patient-isolated CD34+/CD38- AML cells revealed that the levels of anti-apoptotic protein BCL2L12 were downregulated after CD82 depletion by specific shRNA. Western blot analysis indicated that BCL2L12 was aberrantly expressed in patient-isolated AML cells and AML cell lines. Furthermore, CD82 blockade by a specific antibody downregulated BCL2L12 in parallel with de-phosphorylation of STAT5 and AKT, while pharmacological inhibition of STAT5 and AKT activation decreased BCL2L12 expression in leukemia cells. In addition, shRNA-mediated downregulation of BCL2L12 increased the levels of cleaved caspase 3and suppressed proliferation of leukemia cells, impairing their engraftment in immunodeficient mice. Taken together, our results indicate that CD82 regulated BCL2L12 expression via STAT5A and AKT signaling and stimulated proliferation and engrafting of leukemia cells, suggesting that CD82 and BCL2L12 may be promising therapeutic targets in AML.

Dataset Information

Differentiated vs. undifferentiated promyelocytic cell lines (HL-60 and KG-1) and Neutrophiles

Publications

Identification of the tetraspanin CD82 as a new barrier to xenotransplantation.

Similar Datasets

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