Defective NK-cell activation and cytolytic activity in NK cells from a STAT1-deficient patient. (A) Cytometric analysis of STAT1 expression in IL-2–activated NK cells using intracellular staining with anti-STAT1 mAb was performed as described in “Cytokine, ELISA assay, and flow cytometry.” Mean fluorescence intensity is indicated. The results are representative of 3 independent experiments. (B) We performed a flow cytometric analysis of STAT1 phosphorylation (Y701) of IL-2–activated NK cells after treatment with IFN-α (40 U/μL) or medium alone for 20 minutes at 37°C using intracellular staining with an anti-phospho-STAT1-PE mAb. (C) Cytometric analysis of STAT4 phosphorylation in IL-2–activated NK cells using intracellular staining with anti-phospho-STAT4-PE mAb was performed as described in “Cytokine, ELISA assay, and flow cytometry.” Mean fluorescence intensity is indicated. The results are representative of 3 independent experiments. (D) IL-2–activated NK cells were stimulated with IL-12 (200 ng/mL) or IFN-α (104 U/mL) for 24 hours. Supernatants were collected after stimulation, and IFN-γ levels were evaluated by ELISA. (E-F) Defective cytolytic activity of the STAT1-deficient patient against NK-susceptible target cells. Freshly isolated NK cells derived from the patient (▴) or from a healthy donor (■) were tested against the K562 target cells either before (E) or after overnight incubation with rIL-2 (F). The results shown here are representative of 3 independent experiments. Data are mean ± SE. *Significant difference in the response between the patient and the control subject (P < .05).
