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Differential Proteomic Study of Human CD4~+ CD25~+T Cells and CD4~+ CD25~+ T Cells

Author: PengShuangFa
Tutor: WuJun
School: Third Military Medical University
Course: Surgery
Keywords: Regulatory T cells Proteomics Two-dimensional gel electrophoresis Matrix-assisted laser desorption ionization - time of flight mass spectrometry Peptide mass fingerprinting
CLC: R392
Type: PhD thesis
Year: 2003
Downloads: 278
Quote: 0
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Abstract


Body formation and maintenance of immune tolerance mechanism, in addition to the thymic negative selection and peripheral T cell anergy passive mechanisms, studies have shown the initiative mechanism, to cut other cells of the immune response by regulatory T cells in order to maintain immune tolerance. Supplement the cells clear natural CD4 ~ CD25 ~ T cells was first discovered in 1995, occurred after the autoimmune disease and prevent its occurrence has been more and more studies prove the natural CD4 ~ CD25 ~~ T cells with regulatory T cell characteristics , CD4 ~ CD25 ~ T cells to the body of autoreactive T cells and effector T cells selectively suppressing immune homeostasis formation and maintenance of one of the important mechanisms in this cell recently in autoimmunity, transplantation immunity and tumor immunity areas such as more and more attention. Although the cellular functions certainly, but the molecular mechanism of its function is still unclear, such as: the immunomodulatory effects or cytokine pathways through the contact mechanism? CD25 is a hallmark of T cell activation, CD4 ~ CD25 ~ regulatory T cells and on the basis of the immune response after activation of CD4 CD25 to T cells in place different? whether there are more specific than CD25 markers to be found? Differential Proteomics of the post-genomic era for us the whole picture analysis of proteomic expression differences explore the cellular and molecular mechanisms. The two-dimensional gel electrophoresis and matrix-assisted time-of-flight mass spectrometry is used differential proteomics pillar. Functionally related molecular markers and drug targets screening study shows the value of the two kinds of technology. This paper through human CD4 ~ CD25 ~ T cells, and CD4 ~ CD25 ~-T cells in the protein group expression difference analysis as well as differences in protein's identification, in order to explore the CD4 ~ CD25 ~ T CD4 ~ CD25 ~-T cell functional differences in the molecular basis of provide a basis for further screening-specific functionality related protein and intervention targets. First, we apply the bead sorting method from the successful separation of high purity human spleen CD4 ~ CD25 ~ T cells and CD4 ~ CD25 ~-T and its function function test analysis, found that spleen-derived natural CD4 ~ CD25 ~ T cells with low immune response and a significant inhibitory effect on the immune response of CD4 ~ CD25 ~-T. That borne natural human spleen CD4 ~ CD25 ~ T cells with regulatory cell characteristics. With human CD4 ~ CD25 ~-T-cell function varied significantly. We then used two-dimensional gel electrophoresis sorting CD4 ~ CD25 ~ T cells CD4 ~ CD25 ~-T cell protein separation, to obtain a better display of the whole-cell protein gel, through the use of image analysis software digitized quantitative analysis Proteome expression profiling. Proteome of the two kinds of cells in the distribution patterns broadly similar protein spots were concentrated in isoelectric point between pH4 to 8 and a molecular weight ranging from 14kDa to 65kDa average CD4 ~ CD25 to-T-cell plastic Figure 704 point, CD4 ~ CD25 ~ T cells in an average of 685 points. Soft Third Military Medical University PhD On file analysis than found that the expression level of the margin is more than four times the total of 25 protein spots, 17 of them in the higher level of CD4 CD25-T cells express eight points in the CD mine CD25 T cells express higher levels. Finally, we cut 25 protein spots gel digestion, the use of belly mass fingerprint identification strategy successfully identified from 15 points, including 13 of CD4 CD25.T cells, CD4 CD25 T cells 2. Successfully identified proteins, CD4 CD25T cells express higher levels of the protein, including a variety of metabolic enzymes and structural proteins, as well as the signal transduction the protein (LCKBPI), heat shock protein 70 chaperone family members (Bip protein ), signal the molecular lasp a 1, apoptosis-related molecules (casPase to a 7B chain), CD4 10 cD25 T cells express higher levels of the two kinds are nucleic acid metabolism related proteins. These results lay the foundation for further functional studies of related molecules.

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