Construction of a recombinant adenovirus co-expressing truncated human prostate-specific membrane antigen and mouse 4-1BBL genes and its effect on dendritic cells
| Autor: | Heng-cheng Zhu, Zhiyuan Chen, Botao Jiang, Hao Shen, You-lin Kuang, Xiaodong Weng, Xiuheng Liu, Hui Chen |
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| Rok vydání: | 2011 |
| Předmět: |
Cytotoxicity
Immunologic Physiology Apoptosis medicine.disease_cause Dendritic cells Biochemistry law.invention Mice Multiplicity of infection Transduction Genetic law Adenovirus 4-1BBL General Pharmacology Toxicology and Pharmaceutics lcsh:QH301-705.5 lcsh:R5-920 Prostate cancer medicine.diagnostic_test Co-stimulatory molecule General Neuroscience hemic and immune systems General Medicine Flow Cytometry Interleukin-12 Recombinant Proteins Phenotype Recombinant DNA Female lcsh:Medicine (General) Immunology Biophysics Enzyme-Linked Immunosorbent Assay Biology Adenoviridae Flow cytometry Western blot Antigen PSMA medicine Animals Humans CD86 Interleukin-6 Cell Biology Prostate-Specific Antigen Molecular biology Mice Inbred C57BL 4-1BB Ligand HEK293 Cells lcsh:Biology (General) CD80 |
| Zdroj: | Brazilian Journal of Medical and Biological Research v.44 n.3 2011 Brazilian Journal of Medical and Biological Research Associação Brasileira de Divulgação Científica (ABDC) instacron:ABDC Brazilian Journal of Medical and Biological Research, Vol 44, Iss 3, Pp 186-192 (2011) Brazilian Journal of Medical and Biological Research, Volume: 44, Issue: 3, Pages: 186-192, Published: MAR 2011 |
| ISSN: | 0100-879X |
| Popis: | Our aim was to construct a recombinant adenovirus co-expressing truncated human prostate-specific membrane antigen (tPSMA) and mouse 4-1BBL genes and to determine its effect on dendritic cells (DCs) generated from bone marrow suspensions harvested from C57BL/6 mice for which the effect of 4-1BBL on DCs is not clear, especially during DCs processing tumor-associated antigen. Replication deficient adenovirus AdMaxTM Expression System was used to construct recombinant adenovirus Ad-tPSMA-internal ribosome entry site-mouse 4-1BBL (Ad-tPSMA-IRES-m4-1BBL) and Ad-enhanced green fluorescent protein. Day 7 proliferating DC aggregates generated from C57BL/6 mice were collected as immature DCs and further mature DCs were obtained by lipopolysaccharide activated immature DCs. After DCs were exposed to the recombinant adenovirus with 250 multiplicity of infection, the expression of tPSMA and m4-1BBL proteins were detected by Western blot, and the apoptosis and phenotype of DCs were analyzed by flow cytometry. Cytokines (IL-6 and IL-12) in the supernatant were detected by enzyme-linked immunosorbent assay (ELISA). Proliferation of T cells was detected by allogeneic mixed lymphocyte reactions. The tPSMA and m4-1BBL proteins were expressed correctly. The apoptosis rate of DCs transfected with Ad-tPSMA-IRES-m4-1BBL was 14.6%, lower than that of control DCs. The expression of co-stimulatory molecules [CD80 (81.6 ± 5.4%) and CD86 (80.13 ± 2.81%)] up-regulated in Ad-tPSMA-IRES-m4-1BBL-pulsed DCs, and the level of IL-6 (3960.2 ± 50.54 pg/mL) and IL-12 (249.57 ± 12.51 pg/mL) production in Ad-tPSMA-IRES-m4-1BBL-transduced DCs were significantly higher (P < 0.05) than those in control DCs. Ad-tPSMA-IRES-m4-1BBL induced higher T-cell proliferation (OD450 = 0.614 ± 0.018), indicating that this recombinant adenovirus can effectively enhance the activity of DCs. |
| Databáze: | OpenAIRE |
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