| Background: Dendritic cells (DC) are the professional antigen presenting cells. They are the body's sentinels which initiate the strong immune responses to invading microbes, mutant or destroyed cells (possibly also cancer cells), the antigen sources. Immature DC has very active phagocytic function to capture various antigens. The captured antigens in high content are effectively broken down into small peptides. These peptides are loaded onto HLA class I and class II molecules which are abundantly expressed in mature DC having co-stimulatory molecules. These features enable DC to effectively activate T cells and also B cells both in vitro and certainly in vivo. Therefore, the matured human DC expressing a protein which accumulates intracellularly can become a potent vaccine to initiate an immune response to get ride of the antigen source.Goal: Accumulation of p53 protein had been reported in majority of human cancers. Expression of the complete protein instead of using its peptides broadens its clinical use without HLA typing limitation. The present study has mainly focused on how to achieve high expression of p53 in human mature dendritic cells. It might be interesting also to see whether wild type p53 can accumulate in DC without causing apoptosis.Material and Methods:1. To construct and expand a recombinant from a plasmid containing the full length mutant p53 and a retrovirus plasmid pLXSN. Characterize the target gene by PCR & RFLP as well as by DNA sequencing.2. Using p53 expression null Hep3B cell line, transfect p53 gene vector into these cells via lipofection. Identify p53 protein and also hTERT by immuno-cytochemistry and FACS.3. Culture the DC precursors from patients' leukopak or fetal cord blood in AIM V media with SCF, Flt3, GM-CSF, IL-4. Observe the differentiation status of the DC by morphology and FACS.4. Transfect DC in early stage of development with p53 recombinant. Observe its expression in mature DC by immunocytochemistry and FACS.
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