Oligopeptide (ogp, Hiv-1 Tat, [47-57]) Synthesis, Polymerization And Functional Studies And Nrsf / Rest Transcriptional Regulation Research

The thesis consists of two parts. The first part is the synthesis, polymerization ofOsteogenic Growth Peptide (OGP) and HIV-1 tat[47-57], the study of their function;the second part is the study of the effect of neuron restrictive silencer factor/repressorelement-1 silencing transcription factor (NRSF/REST) on gene transcriptionalregulation. The first chapter of part I describes the synergistic effect of OGP with recombinanthuman granulocyte colony stimulating factor (rhG-CSF) on hematopoiesis. In vitroOGP regulated cell proliferation, alkaline phosphatase activity, and matrixmineralization; in vivo OGP increased osteogenesis and blood granulocyte count, enhancedengraftment of bone marrow transplants of mice. We found that sOGP had a remarkablesynergistic action with rhG-CSF on increment of white blood cell number and lymphocytenumber in peripheral blood, so sOGP would be a potential new drug to improve the effectivenessof clinical application of rhG-CSF. The second chapter and third chapter of part I describe design and synthesis ofmacro branched peptides and their application for immunoassay and gene delivery.Two kinds of peptide dendrimers have been used as immunogen, antiviral agents andvehicles for delivery of nucleic acids and drugs. One of these peptide dendrimerscontained poly lysine as the central core to link the peptide chains as the surfacebranching units; another kind of peptide dendrimers contained the multiple antigenpeptide (MAP) system, i.e., containing the peptide chains with only the branchingunits. In this part, a simple method for macro branched peptide synthesis was used. 3Compound containing the double bond between two carbons was introduced intopeptide using reagents such as acryloyl chloride during peptide synthesis, thustransforming to a polymer containing a poly propionyl core matrix with peptidebranches by radical polymerization. Different peptides (proinsulin C peptide, OGP,HIV-1 tat and penetratin) were respectively polymerized by different strategiesaccording to their physical–chemical characteristics. The second chapter describesthe application of poly-C peptide and poly-OGP for antiserum preparation and theapplication for specific immunoassays. The third chapter describes the application ofpoly-tat for plasmid DNA delivery and the result shows that it is a good non-viralvector for gene delivery. The DNA transfection efficiency and cell toxicity are foundrelated with the molecular weight of poly-tat. The second part describes the gene expression variation and cell differentiationafter the knockdown of NRSF/REST. NRSF/REST is one of the negativetranscription factors having the function to make neuronal related gene silencing innon-neuronal cells. In this part, NRSF/REST in HEK 293 cells was knocked downby SiRNA and the influence at gene expression was tested by cDNA microarray. Alot of neuronal related genes were up regulated after knockdown of NRSF/REST andthe variation of gene expression were further testified by real-time PCR. Neuronrestrictive silencer element (NRSE) in up-regulated gene was studied bybioinformatic investigation. The result of neuron specific gene such as synapsin Iexpressed in non-neuronal cells suggested the important role of NRSF/REST at celldifferentiation from non-neuronal cells to neuron.