| Major histocompatibility complex of swine was called swine lymphocyte antigen(SLA), SLA was consist of high polymorphic and closely likage gene cluster,SLA was play a critical role in antigen presentation, immune regulation and disease resistance. The study show that effect of transplantation was significantly different when we selected different haplotype swines as a donor for xenotransplantation.Currently, Wuzhishan minipigs was one of the animal models on xenotransplantation study, the study was aimed to clarify SLA alleles and haplotype distribution of class I and class II on Wuzhishan minipigs, we used molecule cloning sequence and PCR-SSP method to detected SLA haplotype. At the same time, we used complement dependent cytotoxicity tests to analyse relevance of different haplotype individuals with immune rejection in xenotransplantation, we expected to provide some theory basis for xenotransplantation. The major conclusions are as follows:1. We have found 10 alleles on SLA five locus of Wuzhishan minipigs. SLA-1locus have two novel allele sequences, we submit the sequence to Gen Bank and ISAG, meanwhile, we obtained corresponding accession number and allele name,SLA-1*0401wz01(Accession number:KT715501), SLA-1*0703(Accession number:KT194212). SLA-2*1102 was previously reported, but SLA-2*11wz01(Accession number:KT194213) was a novel allele. SLA-3*0401,SLA-3*0403,SLA-DRB1*0901SLA-DRB1*0401 、 SLA-DQB1*0901 and SLA-DQB1*0501 were all previously reported.2. Three SLA class I haplotypes and two class II haplotypes were novel found from the Wuzhishan miniature, which were named and designated as Hp-75.00( SLA-1*0401wz01, SLA-3*0403, SLA-2*11wz01), Hp-76 b.00(SLA-1*0703, SLA-3*0401, SLA-2*11wz01), Hp-76 a.00(SLA-1*0703, SLA-3*0401, SLA-2*1102), Hp-00.27(SLA-DRB1*0901, SLA-DQB1*0901), Hp-00.41(SLA-DRB1*0401, SLA-DQB1*0501).3. Based on 10 alleles sequences imformation obtained from progenitor Wuzhishan minipigs, we established a simple and fast methed to SLA typing forunknown descendant of Wuzhishan minipigs, relying on designed 10 efficient sequence specific primers, we rapidly conduct SLA typing on DNA level for Wuzhishan minipigs and identify four SLA haplotypes present in the founder.4. The homology comprision of vital amino acid residues on HLA binding with the human CD4/CD8 molecule and KIR showed that there had been a highly homology between ten SLA alleles of Wuzhishan minipigs and HLA,simultaneously,amino acid sequence of SLA-DRB1 alleles were consistent with HLA vital amino acid binding with the human CD4 molecule.5. We knowed that the blood type of Wuzhishan minipigs herd was A by the blood typing. the peripheral blood mononuclear cell separated from three hplotype individuals reacted with inactivating human or monkey serum, the result showed that the cell death rate was not significant when the monkey serum reacted with different haplotype individuals, but the cell death rate was significant when the the human serum reacted with different haplotype individuals(p<0.01).The study has firstly systematic completed haplotype typing of SLA classⅠand SLAⅡgenes on wuzhishan minipigs and obtained two homozygous haplotype strains.In vitro studies have shown that the correlation of the immune rejection strength with SLA haplotype type is low, however, the cell death rate was relative low when the PBMC from wuzhishan minipigs which was humanized modified reacted with human or monkey serum. Therefore, Wuzhishan minipigs can be used as one of the good donor for future xenotransplantation. |
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