The impact of rhVWF on FVIII-sensitized CD4⁺T cell proliferation in response to rhF8 restimulation. Splenocytes isolated from rhF8-immunized FVIII^null mice were labeled with CellTrace Violet and cultured with a low (1 U/mL) or a high (10 U/mL) dose of rhF8 with or without 1 U/mL rhVWF for 96 hours. Cells were stained with anti–mouse CD4 and anti–mouse TCRβ antibodies and analyzed by flow cytometry for daughter CD4⁺ T cells. (A) Representative flow cytometry histograms. (B-C) The impact of rhVWF on CD4⁺ T cell proliferation in response to a low dose of rhF8 (B) and a high dose of rhF8 (C). The paired Student t test was used to compare data sets. (D-E) Normalized daughter cell data. The percentage of daughter CD4⁺ T cells in the condition without VWF was defined as 1. The Mann-Whitney U test was used to compare normalized data sets. These data demonstrate that VWF can attenuate FVIII-sensitized CD4⁺ T-cell proliferation in response to rhF8 restimulation.