PMC

Baseline mRNA transcript expressions of tryptase (TPSAB1), chymase (CMA1), and CPA3 are shown. Each LUVA transcript is expressed as mean ± SEM, n=4 experiments, and in parallel are the baseline levels of the same transcripts by LAD2 cells (n=1).

Cells were stimulated with the indicated doses of SCF for 72 hours. Proliferation rates were determined by [³H]thymidine incorporation. Data are mean ± SD, n=3 experiments.

Quantification of β-hexosaminidase (A), PGD₂ (B), TXA₂ (C), and MIP-1β (D) levels in cell supernatants are shown. In some experiments the cells were primed for 3 days with IL-4 (10 ng/mL) before activation. Data are mean ± SD, n= 2 or 3 experiments for LUVA cells, and n=1 or 2 experiments for LAD2 cells.

400× magnifications of LUVA cells are in the left column and of CD34⁺-derived primary MCs are in the right column. Cells stained with toludine blue (A and F), tryptase (B and G), chymase (C and H), cathepsin G (D and I), and CPA3 (E and J) are shown.

Representative flow cytometric analyses are of the expression of c-kit (A), FcγRII/CD32 (B), and FcγRI/CD64 (C). Basal expression of FcεRIα and upregulation of FcεRIα after incubation with IgE (D) or IL-4 (E) are shown. Each analysis was carried out at least 3 times. Isotype controls are shown in gray.

Cells were starved of SCF for 3 hours and then were stimulated for 10 minutes with SCF (100 ng/mL) or medium alone. SDS-PAGE immunoblotting comparing phosphorylated c- kit and total c-kit (A). Phosphorylation of Akt, MEK, p42/44 ERK and CREB in the same experiment (B). The data in a second experiment were similar.

LUVA cells were cultured in medium alone (A), with SCF (B), or with SCF and imatinib mesilate (C). Six week-old primary human cord blood-derived MCs (cbMCs) were cultured with the same conditions (D–F). The percentage of apoptotic cells was measured by surface expression of Annexin V, compared to isotype control in shaded gray. Each analysis was carried out at least 3 times with similar results.