A Single Amino Acid Substitution Prevents Recognition of a Dominant Human Aquaporin-4 Determinant in the Context of HLA-DRB1*03:01 by a Murine TCR

作者：Arellano Benjamine; Hussain Rehana; Miller Little William A; Herndon Emily; Lambracht Washington Doris; Eagar Todd N; Lewis Robert; Healey Don; Vernino Steven; Greenberg Benjamin M; Stueve Olaf

来源：PLos One, 2016, 11(4): e0152720.

DOI：10.1371/journal.pone.0152720

Background
Aquaporin 4 (AQP4) is considered a putative autoantigen in patients with Neuromyelitis optica (NMO), an autoinflammatory disorder of the central nervous system (CNS). HLA haplotype analyses of patients with NMO suggest a positive association with HLA-DRB1*03:01. We previously showed that the human (h) AQP4 peptide 281-300 is the dominant immunogenic determinant of hAQP4 in the context of HLA-DRB1*03:01. This immunogenic peptide stimulates a strong Th-1 and Th-17 immune response. AQP4(281-300)-specific encephalitogenic CD4(+) T cells should initiate CNS inflammation that results in a clinical phenotype in HLA-DRB1*03:01 transgenic mice.
Methods
Controlled study with humanized experimental animals. HLA-DRB1*03:01 transgenic mice were immunized with hAQP4281-300, or whole-length hAQP4 protein emulsified in complete Freund's adjuvant. Humoral immune responses to both antigens were assessed longitudinally. In vivo T cell frequencies were assessed by tetramer staining. Mice were followed clinically, and the anterior visual pathway was tested by pupillometry. CNS tissue was examined histologically post-mortem. Flow cytometry was utilized for MHC binding assays and to immunophenotype T cells, and T cell frequencies were determined by ELISpot assay.
Results
Immunization with hAQP4281-300 resulted in an in vivo expansion of antigen-specific CD4(+) T cells, and an immunoglobulin isotype switch. HLA-DRB1*03:01 TG mice actively immunized with hAQP4(281-300), or with whole-length hAQP4 protein were resistant to developing a neurological disease that resembles NMO. Experimental mice show no histological evidence of CNS inflammation, nor change in pupillary responses. Subsequent analysis reveals that a single amino acid substitution from aspartic acid in hAQP4 to glutamic acid in murine (m)AQP4 at position 290 prevents the recognition of hAQP4(281-300) by the murine T cell receptor (TCR).
Conclusion
Induction of a CNS inflammatory autoimmune disorder by active immunization of HLA-DRB1*03:01 TG mice with human hAQP4281-300 will be complex due to a single amino acid substitution. The pathogenic role of T cells in this disorder remains critical despite these observations.

• 出版日期2016-4-7