Primary cells and culture conditions

BMDDC were generated according to previous reported procedures (61). Bone marrow cells (4–5 × 10⁵/ml, 10 ml/plate) were plated in RPMI-1640 (HyClone, Logan, UT), supplemented with 10% fetal bovine serum (FBS), 100 U/ml penicillin, 100 μg/ml streptomycin (all from Mediatech, Manassas, VA), and 20 ng/ml recombinant mouse granulocyte macrophage colony-stimulating factor (GM-CSF) (BioAbChem, Ladson, SC) (DC medium) into 100-mm culture dishes (Sarstedt, Newton, NC). Fresh DC medium was added on day 4 and was gently replaced by fresh DC medium containing 10 ng/ml recombinant mouse GM-CSF on day 7. Immature BMDDC (nonadherent and loosely adherent cells) were used in experiments on day 8.

Extraction of the MEF cells from Gstp1/p2^+/+ or Gstp1/p2^−/− mice and the establishment of immortalized cultures were described previously (39). Cells were maintained in Eagle's Minimum Essential Medium (EMEM) (ATCC, Manassas, VA) containing 10% FBS, 1% nonessential amino acid, 100 U/ml penicillin, and 100 μg/ml streptomycin.