Figure 8.

Developing Gα_i-deficient transitional B cells in OP9-Delta-like 1 (DL1) cultures poorly upregulate ADAM10 expression. (A) Representative flow cytometry to analyze IL-7 cultured bone marrow cells. Bone marrow cells from Gnai2^fl/fl or double knockout (DKO) mice cultured with IL-7 for 4 days. Following removal of the non-adherent the cells, the remaining cells were immunostained for CD19, CD93, IgM, IgD, CD23, ADAM10, and CD81. Shown are CD19 versus CD93; IgM versus CD23 gated on CD19⁺CD93⁺; IgD versus CD23 gated on CD19⁺CD93⁺; CD81 versus ADAM10 gated on CD19⁺CD93⁺IgM⁺IgD⁻ [transitional B cell type 0 (T0)]; and CD81 versus ADAM10 gated on CD19⁺CD93⁺IgM⁺IgD⁺CD23⁻ [transitional B cell type 1 (T1)]. (B) Representative flow cytometry results to analyze B cell development and ADAM10 expression 2 days after plating on OP9-DL1 cells. Bone marrow cells from panel (A) further cultured on OP9-1 cell in the presence of BAFF. The developing B cells from the and DKO mice were characterized by flow cytometry as in panel (A), but in addition CD81 versus ADAM10 expression is shown on CD19⁺CD93⁺IgM⁺IgD⁺CD23⁺ [transitional B cell type 2 (T2)] cells. (C) Charts of the % of ADAM10⁺CD81⁺ T0–T2 cells in OP9-DL1 cultures of WT and DKO bone marrow (above) or only the T1 cells (below). Results from the analysis of bone marrow prepared from three WT and four DKO mice. Data are mean ± SEM (*p < 0.05, **p < 0.005).