| Xenotransplantation is perceived to be the most logical solution to the increasing shortage of donor organs for clinical transplantation. The major barrier on xenotransplantation is hyperacute rejection(HAR). The presence of natural antibodies and activation of the complement cascade during xenogeneic transplantation lead to the destruction of the transplanted organ within minutes or hours. The a l,3-galactosyle(α -Gal) epitope is the major xenoantigen. This epitope is formed by α 1,3-galactosyltransferase (α -GT), which is present in all mammals except man, apes, and Old Word Monkeys. The expression of a -Gal on endothilium is more than bone, cartilage, muscle and nerve tissue. In humans, a -GT is absent and therefore a -Gal is not expressed. Consequently, anti- a Gal antibodies are acquired naturally resulting from exposure to a -Gal-expressing enteric flora and results in humans hyperacutely rejecting a Gal-positive tissue.Among regenerative therapies, cell transplantation offers a promising approach to restore cellular functions and replace or improve organ function. But directly using human cells may suffer from a limited supply. Difficulties in testing the quality of freshly isolated cells prior to implantation and in guaranteering implant oncosafety, particularly when cells have been submitted to genetic modification, also hamper cell allotransplantation. In addition, the use of human embryonic stem(ES) cells, which constitute a promising multipotent cell source that may be expandable prior to differention, currently remains limited for ethical and technical reason. Therefore, effort directed towords adapting animal cell for transplantation into patients remains a worthwhile approach. When compared with organs, cell transplants offer multiple advantages that could largely facilitate their use for clinical application. In vitro cultured cell sources constitute relatively pure preparations, whose quality can be determined before implantation. The absence of donor professional antigen-presenting and immune cells are believed to decrease their immunogenicity. In most cases, xenogeneic cellular implants derive their blood supply from the ingrowth ofrecipient vessels, and thereby are not exposed to the two major hurdles of hyperacute and acute vascular rejection, mediated by antibodies and complement, that affect the outcome of vascularized xenogeneic organ transplants. Usually cell immunoreaction is occurred on cell xenotransplantation, but now some dates show that humour immunoreaction is also occurred. The study of a -Gal will be overcome humour immunoreaction on cell xenotransplantation.Along with the development on mouse embryonic stem cell system and embryo rebuild, gene targeting is applied in almost biology field. It is an important technology on study protein and gene function. Now lots of gene knockout models have been established.To study the role of a -Gal on xenotransplantation and make a -GT gene targeting model in green fluorescent protein mouse, we detect the distribution of a -Gal on mouse embryonic stem cells. The a -GT gene targeting on mouse embryonic stem cells has been finished. The main results are as follows.1. The expression of a -Gal was determined by affinity immuno-histochemistry and Hochest 33258. The a -Gal remarkably positive staining was appeared in membrane and plasm on ES cell, but nuclear is negative. It indicates that we must pay attention to HAR in cell xenotransplantation, especially differentiation ES cell in vivo.2. The long arm (5.1 kb) and short arm (3.5kb) in a -GT gene was isolated from the genomic DNA of C57B/6J mouse by the long polymerase chain reaction (PCR). These fragments were cloned into ploxP vector(7.5kb) on both side of the neomycin gene. Then, we obtain a gene targeting vector ploxP-GT(16.4kb). The homologous sequence is about 9.0kb and identified by base sequence, enzyme digest and PCR. This method is economical and effective.3. The Not I -linearized targeting vector was electroporated into C57BL/6J mouse ES...
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