title: Prolonged expression of MHC class I - peptide expression in bone marrow derived retrovirus transfected matured dendritic cells by continuous centrifugation in the presence of IL-4.
abstract: OBJECTIVE
  Dendritic cells (DCs) are potent antigen presenting cells which proceed from immature to a mature stage during their differentiation. There are several methods of obtaining long lasting mature antigen expressing DCs and different methods show different levels of antigen expressions. We investigated bone marrow derived DCs for the degree of maturation and genetically engineered antigen presentation in the presence of interleukin-4 (IL-4) as a maturity enhancer.
  
  
  METHODS
  DCs and transfected retrovirus were cultured together in the presence of granulocyte-macrophage colony stimulating factor (GMCSF)-IL4, GMCSF +IL4, lipopolysaccharide (LPS). B 7.1, B7.2 and CD11c were measured by the degree of immune fluorescence using enhanced green fluorescent protein (EGFP) shuttled retrovirus transfected antigen. Degree of MHC class I molecule with antigen presentation of antigen was also evaluated by fluorescence activated cell sorting. The antigen presenting capacity of transfected DCs was investigated. Bone marrow DCs were generated in the presence of GMCSF and IL-4 in vitro. Dividing bone marrow cells were infected with EGFP shuttled retrovirus expressing SSP2 by prolonged centrifugation for three consecutive days from day 5, 6 and 7 and continued to culture in the presence of GMSCF and IL-4 until day 8.
  
  
  RESULTS
  IL-4 as a cytokine increased the maturation of retrovirus transfected DCs by high expression of B 7-1 and B 7-2. Also, IL-4 induced DC enhanced by the prolonged centrifugation and it was shown by increased antigen presentation of these dendric cells as antigen presenting cell (APC). Cytolytic effects were significantly higher in cytotoxic T cell response (CTLs) mixed with transfected DCs than CTLs mixed with pulsed DCs.
  
  
  CONCLUSIONS
  There was an enhanced antigen presentation by prolonged expression of antigen loaded MHC class I receptors in DCs in the presence of IL-4 by prolonged centrifugation.
authors.affiliation: Department of Pharmacology, Faculty of Medicine, University of Ruhuna, Sri Lanka. menik@med.ruh.ac.lk
authors.forename: L M
authors.initials: LM
authors.lastname: Hettihewa
mesh_terms: D017951:Antigen Presentation / Q000187:drug effects / Q000276:immunology; D000938:Antigen-Presenting Cells / Q000276:immunology / Q000378:metabolism; D060887:B7 Antigens / Q000235:genetics / Q000378:metabolism; D001853:Bone Marrow / Q000276:immunology* / Q000378:metabolism; D002454:Cell Differentiation / Q000235:genetics / Q000276:immunology; D002498:Centrifugation; D003713:Dendritic Cells / Q000187:drug effects / Q000276:immunology*; D005786:Gene Expression Regulation / Q000187:drug effects / Q000276:immunology; D005805:Genes, MHC Class I / Q000276:immunology*; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000276:immunology / Q000494:pharmacology*; D006801:Humans; D015847:Interleukin-4 / Q000276:immunology / Q000494:pharmacology*; D008070:Lipopolysaccharides / Q000494:pharmacology; D010455:Peptides / Q000235:genetics / Q000276:immunology; D012190:Retroviridae / Q000235:genetics; D013602:T-Lymphocytes, Cytotoxic / Q000187:drug effects / Q000276:immunology; D014162:Transfection / Q000379:methods
chemical_list: D060887:B7 Antigens; D008070:Lipopolysaccharides; D010455:Peptides; D015847:Interleukin-4; D016178:Granulocyte-Macrophage Colony-Stimulating Factor