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Research of Therapeutic Vaccine Against Chronic Hepatitis B Based on Heat Shock Protein

Author: YangBingFen
Tutor: LiuZhiMin
School: PLA Military Academy of Medical Sciences
Course: Microbiology
Keywords: Heat shock protein 65 HBcAg gene Chronic hepatitis B Therapeutic vaccine
CLC: R392.1
Type: Master's thesis
Year: 2005
Downloads: 96
Quote: 0
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Abstract


Hepatitis B is a serious public health problem, about 20 million people worldwide infected with HBV ,3-4 million people chronically infected with HBV, of which 25% to 40% will eventually die of cirrhosis of the liver and liver cancer. Of chronic hepatitis B virus (hepatitis B virus, HBV) infection is estimated at 130 million, of which more than 30 million is needed to antiviral therapy in patients with chronic active hepatitis B, the annual treatment cost of approximately $ 300-500 billion yuan. Of a breakthrough in the treatment of chronic hepatitis B, as the main means remains to antiviral therapy. With the development of molecular biomedical, therapeutic vaccines from theory to application of a new direction for chronic hepatitis B treatment. The purpose of this project is to develop a heat shock protein-based therapeutic for chronic hepatitis B vaccine. Discovered in recent years, heat shock protein - peptide complexes throughput receptor (CD91)-mediated endocytosis by macrophage phagocytosis, and then through the proteasomal degradation, and eventually enter the endogenous antigen presentation pathway, thereby activating cell immune. We chose the Mycobacterium tuberculosis heat shock protein 65 (heat shock protein 65, HSP65) and Hepatitis B virus core antigen (Hepatitis B virus core antigen, HBcAg) fusion as the treatment of chronic hepatitis B vaccine. HBcAg protein HSP65 can be presented to the MHC-I antigen presentation pathway, thereby activating cell-mediated immunity. The study showed that the HBcAg is an immunodominant antigen in a self-limited HBV infections strongly HBcAg-specific Th cell response, to generate the corresponding antibodies and T cell epitope of HBcAg to stimulate specific CTL response can be induced in a mouse model. We use HBcAg as a target antigen, its integration in the C-terminus of HSP65 to stimulate specific cellular immunity against HBV. The soluble high expression of genes in E. coli obtained by the HSPHBcAg. For ease of purification, the C-terminal fusion the polar domain Z-tag. A purity exceeding 80% of the samples obtained after the HSPHBcAgZtag protein was purified by ammonium sulfate precipitation, cation exchange chromatography and gel filtration chromatography. The purified fusion protein as the treatment of chronic hepatitis B vaccine. The application of this fusion protein was used to immunize mice, isolated from mouse spleen lymphocytes, the detection of cell proliferative response of T cells in ConA, HBcAg, the results showed that the fusion protein group of T cell proliferation, and memory responses for the presence of HBcAg. The immunized mouse spleen lymphocytes as effector cells, the mouse cell lines were incubated with HBcAg P815 target cells, destruction experimental results show that the fusion protein group than HBcAg group of anti-high. These results lay the foundation for the treatment of hepatitis B vaccine after further study.

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