Figure 5

Suppression of Bim expression in ALPS type IV patient causes resistance to TCR-induced death. (A) Activated human PBL from normal control donors (NC1, NC2), or 6 ALPS type Ia patients were restimulated with increasing doses of OKT3 for 24 h. Percent cell loss was calculated in triplicate by PI exclusion. (B) Activated human PBL from an ALPS type Ib patient or a normal control donor (NC) were treated as in (A). Percent cell loss was calculated in triplicate by PI exclusion. (C) Activated human PBL from normal control donors (NC1, NC2), or 6 ALPS type Ia patients restimulated with 100 ng/ml OKT3 for 0 (-) or 8 h (+), lysed and immunoblotted for BIM. β-actin serves as a loading control. Spot densitometry analysis of the ratio of BIM (EL isoform) to β-actin (normalized to NC1 untreated, dashed line) is plotted below. (D) Activated human PBL from normal control donors (NC1, NC2), an ALPS type Ia patient, and an ALPS Type IV patient (P58) were restimulated with OKT3 for 24 h. Percent cell loss was calculated in triplicate by PI exclusion. Differences in apoptosis sensitivity (relative to NC1 or NC2) were statistically significant (p < 0.01). (E) Activated human PBL as in (D) were restimulated with 100 ng/ml OKT3 for 0 (-) or 8 h (+), lysed and immunoblotted for BIM. β-actin serves as a loading control.