Antibodies to HLA-E in Nonalloimmunized Males: Pattern of HLA-Ia Reactivity of Anti–HLA-E–Positive Sera

Abstract

Natural anti-HLA Abs found in sera of healthy nonalloimmunized males recognize HLA-Ia alleles parallel to those recognized by anti–HLA-E mAbs (MEM-E/02/06/07). Therefore, some of the HLA-Ia Abs seen in healthy males could be due to anti–HLA-E Abs cross-reacting with HLA-Ia. If anti–HLA-E Abs occur in healthy nonalloimmunized males, it can be assessed whether they evoke HLA-Ia reactivity as do mouse HLA-E mAbs. IgG and IgM Abs to HLA-E and HLA-Ia alleles are identified in sera of healthy males using microbeads coated with recombinant denatured HLA-E or a panel of rHLA-Ia alleles. The pattern of allelic recognition is comparable to that of anti–HLA-E mAbs. Sixty-six percent of the sera with HLA-E IgG have a high level of HLA-Ia IgG, whereas 70% of those with no anti–HLA-E Abs have no HLA-Ia Abs. HLA-E IgM/IgG ratios of sera are divided into four groups: IgMLow/IgGLow, IgMHigh/IgGLow, IgMHigh/IgGHigh, and IgMLow/IgGHigh. These groups correspond to anti–HLA-Ia IgM/IgG ratio groups. When HLA-E IgM and IgG are absent or present in males, the IgM or IgG of HLA-Ia are similarly absent or present. The mean fluorescent intensity of HLA-Ia Abs correlates with that of anti–HLA-E Abs. Most importantly, HLA-E and HLA-Ia reactivities of the sera are inhibited by the shared, but cryptic, peptide sequences 117AYDGKDY123 and 137DTAAQIS143. Therefore, Abs to the H chain of HLA-E may be responsible for some of the HLA-Ia allele reactivity of the natural HLA-Ia Ab in human sera. Absence of any anti–HLA-Ia Abs in 112 nonvegans and the presence of the same in vegans suggest that dietary meat proteins might not have induced the natural allo-HLA Abs.

Therapeutic preparations of IVIg contain naturally occurring anti–HLA-E antibodies that react with HLA-Ia (HLA-A/-B/-Cw) alleles

“…The recombinant HLA-E, HLA-F, and HLA-G heavy chains are individually attached by a process of simple chemical coupling to 5.6-mm polystyrene microspheres, which are internally dyed (by One L) with infrared fluorophores. [13][14][15] The HLA-Ia microbeads have built-in control beads: positive beads that are coated with human IgG (or murine IgG when mAb was used) and negative beads that are coated with serum albumin (HSA/BSA). For HLA-E, HLA-F, and HLA-G, the control beads (both positive and negative) were added separately.…”

“…Using dual-laser flow cytometry (Luminex xMAP multiplex technology), the single antigen assays were performed for data acquisition and analysis of anti-HLA-Ia and anti-HLA-E Abs. [13][14][15] The LABScreen Single Antigen (One L) assay consists of a panel of color-coded microspheres (single antigen beads) coated with HLA antigens to identify antibody specificities. The single recombinant HLA-Ia antigens in LS1A04-lot 003, 004, and 005 were used for screening IVIg and human standard IgG.…”

“…19 Immunoassay with single-HLA antigen-coated microbeads To detect IgG reactivity to HLA-E and HLA-Ia alleles in IVIg/purified human IgG/murine mAbs, multiplex Luminex-based immunoassay (One L, Inc) was used, as described elsewhere. [13][14][15] IVIg and standard human IgG from commercial sources were serially diluted (from 1/2, ending in 1/256) with phosphate-buffered saline (PBS; pH 7.2) and added to the wells containing the antigen-coated microbeads. Anti-HLA-E mAbs were developed in-house (vide infra), and the supernatants collected from the Figure 1.…”

“…Consequently, it may contain HLA antibodies because several allo-HLA antibodies are known to occur naturally in the sera of both men (nonalloimmunized) and women. [9][10][11][12][13] MoralesBuenrostro et al 11 demonstrated the specificities and incidence of these allo-HLA-I Abs in nonalloimmunized males. Neither the origin nor cause of anti-HLA Abs occurring in healthy individuals has been established.…”

“…14,15 These observations were extended to the sera of nonalloimmunized males to document that the observed HLA-Ia reactivity in human sera could be due to anti-HLA-E Abs. 13 To ascertain whether immunizing humans with HLA-E would augment anti-HLA-E Abs with HLA-Ia reactivity, we examined sera of melanoma patients immunized with autologous melanoma cells grown in interferon-g (IFN-g), a cytokine known to induce overexpression of HLA-E. [16][17][18] The immunized patients not only showed the augmentation of anti-HLA-E Abs, confirming the immunogenicity of HLA-E in humans, but also showed increased reactivity with HLA-Ia (single antigen) coated onto microbeads. 18 Indeed, the pattern of HLA-Ia reactivity in immunized patients corresponded with the pattern of HLA-Ia reactivities exhibited by some of the anti-HLA-E mAbs.…”