Development of a humanized HLA- A30 transgenic mouse model

Meng-min Zhu; Bo-wen Niu; Ling-ling Liu; Hua Yang; Bo-yin Qin; Xiu-hua Peng; Li-xiang Chen; Yang Liu; Chao Wang; Xiao-nan Ren; Chun-hua Xu; Xiao-hui Zhou; Feng Li

doi:10.1002/ame2.12225

摘要：Background : There are remarkable genetic differences between animal major histo-compatibility complex (MHC) systems and the human leukocyte antigen (HLA) sys-tem. HLA transgenic humanized mouse model systems offer a much better method to study the HLA- A- related principal mechanisms for vaccine development and HLA- A- restricted responses against infection in human. Methods : A recombinant gene encoding the chimeric HLA- A30 monochain was con-structed. This HHD molecule contains the following: α1- α2 domains of HLA- A30, α3 and cytoplasmic domains of H- 2D b , linked at its N- terminus to the C- terminus of human β2m by a 15- amino- acid peptide linker. The recombinant gene encoding the chimeric HLA- A30 monochain cassette was introduced into bacterial artificial chro-mosome (BAC) CH502- 67J3 containing the HLA- A01 gene locus by Red- mediated homologous recombination. Modified BAC CH502- 67J3 was microinjected into the pronuclei of wild- type mouse oocytes. This humanized mouse model was further used to assess the immune responses against influenza A virus (H1N1) pdm09 clinically isolated from human patients. Immune cell population, cytokine production, and his-topathology in the lung were analyzed. Results : We describe a novel human β2m- HLA- A30 (α1α2)- H- 2D b (α3 transmembrane cytoplasmic) (HHD) monochain transgenic mouse strain, which contains the intact HLA- A01 gene locus including 49 kb 5?- UTR and 74 kb 3?- UTR of HLA- A01*01. Five transgenic lines integrated into the large genomic region of HLA- A gene locus were obtained, and the robust expression of exogenous transgene was detected in various tissues from A30- 18# and A30- 19# lines encompassing the intact flanking sequences. Flow cytometry revealed that the introduction of a large genomic region in HLA- A gene locus can influence the immune cell constitution in humanized mice. Pdm09 in-fection caused a similar immune response among HLA- A30 Tg humanized mice and wild- type mice, and induced the rapid increase of cytokines, including IFN- γ, TNF- α, and IL- 6, in both HLA- A30 humanized Tg mice and wild- type mice. The expression of HLA- A30 transgene was dramatically promoted in tissues from A30- 9# line at 3 days post-i nfection (dpi). Conclusions : We established a promising preclinical research animal model of HLA- A30 Tg humanized mouse, which could accelerate the identification of novel HLA- A30- restricted epitopes and vaccine development, and support the study of HLA- A- restricted responses against infection in humans.