Background:Human leukocyte antigen(HLA)-DP is much less studied than other HLA class Ⅱ antigens,that is,HLA-DR and HLA-DQ,etc.However,the accumulating data have suggested the important roles of DP-restricted response...Background:Human leukocyte antigen(HLA)-DP is much less studied than other HLA class Ⅱ antigens,that is,HLA-DR and HLA-DQ,etc.However,the accumulating data have suggested the important roles of DP-restricted responses in the context of cancer,allergy,and infectious disease.Lack of animal models expressing these genes as authentic cis-haplotypes blocks our understanding for the role of HLA-DP haplotypes in immunity.Methods:To explore the potential cis-acting control elements involved in the tran-scriptional regulation of the HLA-DPA1/DPB1 gene,we performed the expression analysis using bacterial artificial chromosome(BAC)-based transgenic humanized mice in the C57BL/6 background,which carried the entire HLA-DP401 gene locus.We further developed a mouse model of Staphylococcus aureus pneumonia in HLA-DP401 humanized transgenic mice,and performed the analysis on the expres-sion pattern of HLA-DP401 and immunological responses in the model.Results:In this study,we screened and identified a BAC clone spanning the entire HLA-DP gene locus.DNA from this clone was analyzed for integrity by pulsed-field gel electrophoresis and then microinjected into fertilized mouse oocytes to produce transgenic founder animals.Nine sets of PCR primers for regional markers with an average distance of 15 kb between each primer were used to confirm the integrity of the transgene in the five transgenic lines carrying the HLA-DPA1/DPB1 gene.Transgene copy numbers were determined by real-time PCR analysis.HLA-DP401 gene expression was analyzed at the mRNA and protein level.Although infection with S aureus Newman did not alter the percentage of immune cells in the spleen and thymus from the HLA-DP401-H2-Aβ1 humanized mice.Increased expression of HLA-DP401 was observed in the thymus of the humanized mice infected by S aureus.Conclusions:We generated several BAC transgenic mice,and analyzed the expres-sion of HLA-DPA1/DPB1 in those mice.A model of S aureus-induced pneumonia in the HLA-DP401-H2-Aβ1^(-/-)humanized mice was further developed,and S aureus in-fection upregulated the HLA-DP401 expression in thymus of those humanized mice.These findings demonstrate the potential of those HLA-DPA1/DPB1 transgenic humanized mice for developing animal models of infectious diseases and MHC-associated immunological diseases.展开更多
基金National Science and Technology Major Project,Grant/Award Number:2017ZX10304402-001-006 and 2017ZX10304402-001-012Shanghai Professional Platform for High-level Biosafety Pathogenic Microorganism Detection,Grant/Award Number:18DZ2293000+1 种基金Scientific Research Projects of Shanghai Science and Technology Commission,Grant/Award Number:19140905300Shanghai Public Health Clinical Center General Program and the Start-on Funding,Grant/Award Number:KY-GW-2017-06,KY-GW-2018-11,KY-GW-2018-04,KY-GW-2019-11 and KY-GW-2019-19。
文摘Background:Human leukocyte antigen(HLA)-DP is much less studied than other HLA class Ⅱ antigens,that is,HLA-DR and HLA-DQ,etc.However,the accumulating data have suggested the important roles of DP-restricted responses in the context of cancer,allergy,and infectious disease.Lack of animal models expressing these genes as authentic cis-haplotypes blocks our understanding for the role of HLA-DP haplotypes in immunity.Methods:To explore the potential cis-acting control elements involved in the tran-scriptional regulation of the HLA-DPA1/DPB1 gene,we performed the expression analysis using bacterial artificial chromosome(BAC)-based transgenic humanized mice in the C57BL/6 background,which carried the entire HLA-DP401 gene locus.We further developed a mouse model of Staphylococcus aureus pneumonia in HLA-DP401 humanized transgenic mice,and performed the analysis on the expres-sion pattern of HLA-DP401 and immunological responses in the model.Results:In this study,we screened and identified a BAC clone spanning the entire HLA-DP gene locus.DNA from this clone was analyzed for integrity by pulsed-field gel electrophoresis and then microinjected into fertilized mouse oocytes to produce transgenic founder animals.Nine sets of PCR primers for regional markers with an average distance of 15 kb between each primer were used to confirm the integrity of the transgene in the five transgenic lines carrying the HLA-DPA1/DPB1 gene.Transgene copy numbers were determined by real-time PCR analysis.HLA-DP401 gene expression was analyzed at the mRNA and protein level.Although infection with S aureus Newman did not alter the percentage of immune cells in the spleen and thymus from the HLA-DP401-H2-Aβ1 humanized mice.Increased expression of HLA-DP401 was observed in the thymus of the humanized mice infected by S aureus.Conclusions:We generated several BAC transgenic mice,and analyzed the expres-sion of HLA-DPA1/DPB1 in those mice.A model of S aureus-induced pneumonia in the HLA-DP401-H2-Aβ1^(-/-)humanized mice was further developed,and S aureus in-fection upregulated the HLA-DP401 expression in thymus of those humanized mice.These findings demonstrate the potential of those HLA-DPA1/DPB1 transgenic humanized mice for developing animal models of infectious diseases and MHC-associated immunological diseases.
