Use of human antigen presenting cell gene array profiling to examine the effect of human T-cell leukemia virus type 1 Tax on primary human dendritic cells
Department of Microbiology and Immunology, Institute for Molecular Medicine and Infectious Disease, Center for Molecular Virology and Neuroimmunology, Center for Cancer Biology, Drexel University College of Medicine, Philadelphia, PA 19102, USA. Journal of NeuroVirology
(Impact Factor: 2.6).
03/2006; 12(1):47-59. DOI: 10.1080/13550280600614981
Human T-cell leukemia virus type 1 (HTLV-1) is etiologically linked to adult T-cell leukemia and a progressive demyelinating disorder termed HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP). One of the most striking features of the immune response in HAM/TSP centers on the expansion of HTLV-1-specific CD8(+) cytotoxic T lymphocyte (CTL) compartment in the peripheral blood and cerebrospinal fluid. More than 90% of the HTLV-1-specific CTLs are directed against the viral Tax (11-19) peptide implying that Tax is available for immune recognition by antigen presenting cells, such as dendritic cells (DCs). DCs obtained from HAM/TSP patients have been shown to be infected with HTLV-1 and exhibit rapid maturation. Therefore, we hypothesized that presentation of Tax peptides by activated DCs to naIve CD8(+) T cells may play an important role in the induction of a Tax-specific CTL response and neurologic dysfunction. In this study, a pathway-specific antigen presenting cell gene array was used to study transcriptional changes induced by exposure of monocyte-derived DCs to extracellular HTLV-1 Tax protein. Approximately 100 genes were differentially expressed including genes encoding toll-like receptors, cell surface receptors, proteins involved in antigen uptake and presentation and adhesion molecules. The differential regulation of chemokines and cytokines characteristic of functional DC activation was also observed by the gene array analyses. Furthermore, the expression pattern of signal transduction genes was also significantly altered. These results have suggested that Tax-mediated DC gene regulation might play a critical role in cellular activation and the mechanisms resulting in HTLV-1-induced disease.
Available from: Brian Wigdahl
- "Briefly, CD4 + and CD8 + T cells were purified from monocyte depleted PBMCs by using EasySep negative selection enrichment cocktail (StemCell Technologies, Vancouver, Canada). Dendritic cells (DCs) were differentiated from highly purified CD14 + monocytes as described (Ahuja et al., 2006) and maintained in complete RPMI medium supplemented with recombinant human granulocyte macrophagecolony stimulating factor (GM-CSF, 50 ng/ml, PeproTech; Rocky Hill, NJ) and interleukin-4 (IL-4, 10 ng/ml, PeproTech). "
[Show abstract] [Hide abstract]
ABSTRACT: Quantum dots (QDots) are fluorescent semiconductor nanocrystals with a narrow emission spectrum, high quantum yield, and excellent photostability. These unique properties of QDots have been utilized to develop a fluorescent binding assay using biotinylated human T cell leukemia virus type 1 (biot-HTLV-1) conjugated with streptavidin-coated QDots that enabled both qualitative and quantitative analyses of viral binding. The specificity and linearity of the assay was demonstrated utilizing T cells, the primary HTLV-1-susceptible cell population. Furthermore, differential binding of HTLV-1 was analyzed in additional cell types of clinical relevance including primary CD4(+) and CD8(+) T cells, dendritic cells (DCs), monocytes, bone marrow progenitor cells, and epithelial cells. DCs exhibited maximum binding affinity when compared to other examined cell types except the Jurkat and SUP-T1 T cell lines. Finally, blocking antibodies directed against a putative HTLV-1 receptor on DCs; DC-SIGN (dendritic cell-specific ICAM-3-grabbing non-integrin), were utilized to study the inhibition of HTLV-1 binding to target cells. Overall, these results demonstrated that this novel high throughput assay can be utilized to study the binding of a biotinylated virus and has implications for screening of viral binding inhibitors as well as host membrane proteins that may serve as receptors for viral entry.
Journal of Virological Methods 06/2007; 141(2):125-32. DOI:10.1016/j.jviromet.2006.11.043 · 1.78 Impact Factor
Available from: Brian Wigdahl
[Show abstract] [Hide abstract]
ABSTRACT: Human T cell leukemia virus type 1 (HTLV-1) has previously been shown to infect antigen-presenting cells and their precursors in vivo. However, the role these important cell populations play in the pathogenesis of HTLV-1-associated myelopathy/tropical spastic paraparesis or adult T cell leukemia remains unresolved. To better understand how HTLV-1 infection of these important cell populations may potentially impact disease progression, the regulation of HTLV-1 viral gene expression in established monocytic cell lines was examined. U-937 promonocytic cells transiently transfected with a HTLV-1 long-terminal repeat (LTR) luciferase construct were treated with phorbol 12-myristate 13-acetate (PMA) to induce cellular differentiation. PMA-induced cellular differentiation resulted in activation of basal and Tax-mediated transactivation of the HTLV-1 LTR. In addition, electrophoretic mobility shift analyses demonstrated that PMA-induced cellular differentiation induced DNA-binding activity of cellular transcription factors to Tax-responsive element 1 (TRE-1) repeat II. Supershift analyses revealed that factors belonging to the activator protein 1 (AP-1) family of basic region/leucine zipper proteins (Fra-1, Fra-2, JunB, and JunD) were induced to bind to TRE-1 repeat II during cellular differentiation. Inhibition of AP-1 DNA-binding activity by overexpression of a dominant-negative c-Fos mutant (A-Fos) in transient expression analyses resulted in severely decreased levels of HTLV-1 LTR activation in PMA-induced U-937 cells. These results have suggested that following infection of peripheral blood monocytes, HTLV-1 viral gene expression may become up-regulated by AP-1 during differentiation into macrophages or dendritic cells.
Journal of Leukocyte Biology 10/2006; 80(3):640-50. DOI:10.1189/jlb.1205723 · 4.29 Impact Factor
Frontiers in Bioscience 01/2007; 12(8-12):4315. DOI:10.2741/2390 · 3.52 Impact Factor
Data provided are for informational purposes only. Although carefully collected, accuracy cannot be guaranteed. The impact factor represents a rough estimation of the journal's impact factor and does not reflect the actual current impact factor. Publisher conditions are provided by RoMEO. Differing provisions from the publisher's actual policy or licence agreement may be applicable.