Haematopoietic depletion in vaccine-induced neonatal pancytopenia depends on both the titre and specificity of alloantibody and levels of MHC I expression

Abstract

Bovine Neonatal Pancytopenia (BNP) is a disease of calves characterised by haematopoietic depletion, mediated by ingestion of alloantibodies in colostrum. It has been linked epidemiologically to vaccination of the dams of affected calves with a particular vaccine (Pregsure) containing a novel adjuvant. Evidence suggests that BNP-alloantibodies are directed against MHC I molecules, induced by contaminant bovine cellular material from Madin-Darby Bovine Kidney (MDBK) cells used in the vaccine's production. We aimed to investigate the specificity of BNP-alloantibody for bovine MHC I alleles, particularly those expressed by MDBK cells, and whether depletion of particular cell types is due to differential MHC I expression levels. A complement-mediated cytotoxicity assay was used to assess functional serum alloantibody titres in BNP-dams, Pregsure-vaccinated dams with healthy calves, cows vaccinated with an alternative product and unvaccinated controls. Alloantibody specificity was investigated using transfected mouse lines expressing the individual MHC I alleles identified from MDBK cells and MHC I-defined bovine leukocyte lines. All BNP-dams and 50% of Pregsure-vaccinated cows were shown to have MDBK-MHC I specific alloantibodies, which cross-reacted to varying degrees with other MHC I genotypes. MHC I expression levels on different blood cell types, assessed by flow cytometry, were found to correlate with levels of alloantibody-mediated damage in vitro and in vivo. Alloantibody-killed bone marrow cells were shown to express higher levels of MHC I than undamaged cells. The results provide evidence that MHC I-specific alloantibodies play a dominant role in the pathogenesis of BNP. Haematopoietic depletion was shown to be dependent on the titre and specificity of alloantibody produced by individual cows and the density of surface MHC I expression by different cell types. Collectively, the results support the hypothesis that MHC I molecules originating from MDBK cells used in vaccine production, coupled with a powerful adjuvant, are responsible for the generation of pathogenic alloantibodies.

The risks of using allogeneic cell lines for vaccine production: the example of Bovine Neonatal Pancytopenia

“…Subsequently this was supported by the finding that Pregsure© vaccinated cows generate a unique profile of MHC I-specific alloantibodies, presumably reflecting the degree of alloantigen mismatch between their MHC I alleles and those of the vaccine cell line [11,12]. However, the situation in calves appears to be more complex with BNP-alloantibodies also showing varying degrees of cross reactivity with MHC I alleles not present on MDBK cells [11,12]. This is consistent with the nature of MHC I sequence polymorphism, where short stretches of amino acids are frequently shared by different alleles.…”

“…Using transfected cell lines, sera from BNP cows were also shown to bind specifically to the MHC I alleles present on MDBK cells and to induce complement-mediated cytotoxicity [11]. Furthermore, in vitro experiments showed that BNP alloantibodies preferentially target cells with high MHC I expression and that this correlates with the hematopoietic cell types affected in BNP [11,12,16] (Figure 2 and Section 2). Using a modified MDBK cell line with no MHC I expression (beta-2-microglobulin knockout) it was confirmed that MHC I is the dominant target of BNP alloantibodies and depending on the BNP dam accounts for 46-91% of the alloantibody specificity [12].…”

“…It is now clear that BNP is caused by vaccine-induced maternal alloantibodies which are transferred from mother to calf via the colostrum (first milk rich in maternal antibodies). These alloantibodies, which are specific for major histocompatibility complex class I (MHC I) molecules, are generated in response to residual cell-line material found within a specific vaccine containing a novel adjuvant formulation [8][9][10][11][12]. BNP therefore highlights important issues regarding the safety of using allogeneic cell lines for vaccine production.…”

“…Furthermore, serum and colostrum from BNP cows were shown to contain alloantibodies that bind to peripheral blood leukocytes, platelets and bone marrow cells [9,11,12,[18][19][20][21], and intravenous infusion of these antibodies was sufficient to induce the disease [9]. The bovine epitheliochorial placenta presents a complete barrier to the transfer of immunoglobulins from the cow to the fetus, and thus the calf is completely dependent on ingestion of colostrum for maternal immunoglobulin transfer [22], explaining why BNP only occurs postpartum.…”

“…Following identification of MHC I as the antigenic target in BNP, it was hypothesized that epitope differences between the MHC I alleles expressed by MDBK cells and a given cow would determine the induction of alloantibodies and, conversely, that epitope similarities between MDBK cells and a given calf would determine its propensity to develop the disease [8,9]. Subsequently this was supported by the finding that Pregsure© vaccinated cows generate a unique profile of MHC I-specific alloantibodies, presumably reflecting the degree of alloantigen mismatch between their MHC I alleles and those of the vaccine cell line [11,12]. However, the situation in calves appears to be more complex with BNP-alloantibodies also showing varying degrees of cross reactivity with MHC I alleles not present on MDBK cells [11,12].…”