Immunoglobulin isotypes and functional anti-FVIII antibodies in response to FVIII treatment in Balb/c and C57BL/6 haemophilia A mice

Abstract

Previous studies have demonstrated that genetic factors play an important role in determining the likelihood of formation of anti-factor VIII (FVIII) antibodies in haemophilia A patients. We were interested in characterizing the spectrum of FVIII antibody formation and the primary and secondary immune responses after FVIII administration in two different exon 16-disrupted haemophilia A mouse strains, Balb/c and C57BL/6. Balb/c and C57BL/6 E16 haemophilia A mice were used in all experiments. Total FVIII antibodies and FVIII inhibitors were measured using ELISA and Bethesda assays respectively. T- and B-cell cytokines were quantified using ELISA and flow cytometry. FVIII antibodies, but not functional inhibitors were detectable 1 week after the first FVIII treatment in both strains. These antibodies mainly belonged to the IgM and IgA isotypes. After the fourth FVIII treatment, neutralizing anti-FVIII antibodies were detected in both mouse strains: Balb/c (mean inhibitory titer 58 BU) and C57BL/6 (mean inhibitory titer 82 BU). IgG1 levels were similar in both strains but the IgG2A and IgG2B subclasses were higher in C57BL/6 mice. The results of intracellular cytokine staining of T cells indicated that the FVIII-treated C57BL/6 mice produced more IL10 and Th1 cytokines than the FVIII-treated Balb/c mice. These studies show that C57BL/6 mice develop a stronger immune response towards FVIII than Balb/c mice. We propose that the enhanced Th1 and IL10 cytokine micro-environment induced in C57BL/6 mice is responsible for this difference. Therefore, genetic strain-dependent differences must be considered when evaluating immunological outcomes in mouse models of haemophilia A.

Adjuvant facilitates tolerance induction to factor VIII in hemophilic mice through a Foxp3-independent mechanism that relies on IL-10

“…Our findings on the different isotype preferences by the 2 strains of HemA mice are consistent with previous reports. 29 We confirmed tolerance induction could also be achieved in HemA mice when hFVIII-alum was administered subcutaneously and the anti-CD4 mAb intravenously ( Figure 3F). …”

Concurrent influenza vaccination reduces anti-FVIII antibody responses in murine hemophilia A

“…25,26 Furthermore, the adjuvanted-vaccine modulation of the anti-FVIII response with the associated increase in plasma levels of FVIII-specific IgG1 suggest that the addition of the M59 adjuvant alters the Th profile to a predominately Th2 response, a role that has previously been suggested. 27,28 However, these results may be strain specific as it has For personal use only. on May 9, 2018. by guest www.bloodjournal.org From been suggested that the role of MF59 is to simply augment the default immune profile rather than biasing the profile.…”

Minimal modification in the factor VIII B-domain sequence ameliorates the murine hemophilia A phenotype

“…31,32 To date, there is no conclusive evidence on distinct rates of inhibitor formation comparing recombinant BDD or full-length FVIII. 8 Moreover, the B-domain sequence is the most polymorphic of all the FVIII domains.…”