腺相关病毒介导PSA基因转染DC的实验研究ppt课件

1、腺相关病毒介导腺相关病毒介导PSAPSA基因基因转染转染DCDC的实验研讨的实验研讨尤长宣尤长宣 罗荣城罗荣城 梁卫江梁卫江南方医科大学南方医院肿瘤中心南方医科大学南方医院肿瘤中心The Oncology Department of The Oncology Department of Nanfang Hospital, Nanfang Hospital, the Southern Medical the Southern Medical University, Guangzhou University, Guangzhou Skin & Mucous Membranesrapidly

2、 regenerating surfaces, peristaltic movement, mucociliary escalator, vomiting, flow of urine/tears, coughingCellular and Humoral Defenseslysozyme, sebaceous/mucous secretions, stomach acid, commensal organisms, complement proteins, phagocytosis, natural killer cellsInvasion& infectionBarriersInn

3、ate immunityAdaptive immunityInflammationCellular and Humoral DefensesAntibodies, cytokines, helper T cells, cytotoxic T cellsCTLMK-0069-01Antigen presentationTumor antigenAntigen positive tumor cellDendritic cells express a variety of cell surface molecules which play key roles in their interaction

4、s with T cells. The MHC molecules present antigenic fragments (peptides) to T cells in a form that can be recognized in a highly specific fashion by the appropriate T cell receptor, which is unique for every T cell and every antigen. The costimulatory molecules (CD80 and CD86) provide a second-signa

5、l that amplifies the antigen recognition process. Accessory molecules like ICAMs and LFAs enhance the physical interaction between T cells and DCs. Dendritic Cell Properties Disadvantages of Protein/peptide loadingT1/2 of proteins/peptides is very short. All are rapidly lost.2) Most protocols for ge

6、nerating cytotoxic T lymphocytesCTL take 3-5 weeks.Vectors for Gene deliveryRetroviruses (Retrov) (8kb, RNA, enveloped) Adenoviruses (Ad) (35kb, DNA, non- enveloped) Adeno-Associated Virus (AAV) (5kb, DNA, non- enveloped) Retroviruses (Retrov) (8kb, RNA, enveloped) 1) Transgene inactivation by DNA m

7、ethylation2) Causes cancer: proviral chromosomal integration activates proto-oncogene 3) Retrovirus vectors are easily destroyed, easily inactivated.Adenoviruses (Ad) (35kb, DNA, non-enveloped)-are highly immunogenic, causing inflammation and cell death，that is the non-therapeutic immune response-ca

8、n not infect non-dividing cellsAdeno-Associated Virus (AAV) (5kb, DNA, non-enveloped) non-pathogenic low immune response chromosomal integration long term expression in vivo infect dividing and non-dividing cells Overview of Immune FunctionsrAAV/ PSAHarvestnon-adherentCellsNon adherentCells + IL-2Incubate2-4hr, 37oC AdherentCellsLymphocytes 0.5x107/3ml per wellBloodFicollIsopaqueDilute plasma lymphocytesRed cells granulocytesplateletsrAAV infection Chromium release assayprimingday 0 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15GM-CSFIL-4IL-2 and IL-7Mixed with T-cells or