Figure 3.

Analysis of cell death in SLE patients and healthy controls in response to stimulation with H₂O₂. PBLs from 15 SLE patients and 15 controls were cultured in the presence and absence of 50 μM H₂O₂ for 16 hours and analyzed by fluorescence microscopy and flow cytometry. A, Fluorescence micrographs of H₂O₂-stimulated PBLs from representative control and SLE patient donors. Cells were stained with annexin V–fluorescein isothiocyanate (FITC) and propidium iodide (PI). After 16 hours' incubation in the absence of H₂O₂ (no treatment), the frequency of annexin V–FITC–staining apoptotic cells was increased in SLE PBLs. After 16 hours of treatment with H₂O₂, the frequency of apoptotic cells was lower in SLE PBLs, while that of necrotic cells with swollen nuclei directly staining with PI was elevated (original magnification × 400). B, Flow cytometric analysis of apoptotic (annexin V–positive and PI-negative; lower right quadrant) and necrotic (annexin V–positive and PI-positive; upper right quadrant) cells after stimulation with 50 μM H₂O₂ for 16 hours. PBLs were from different donors than those used in A. C, Apoptosis rates were determined in 15 healthy donors and 15 SLE patients, based on the percentage of annexin V–phycoerythrin (PE)–positive/PI-negative cells. D, After H₂O₂ treatment, necrosis was prominent in SLE patients compared with healthy controls. Data points correspond to the percentage of PI-positive cells within the annexin V–positive population induced by H₂O₂ treatment. See Figure 1 for other definitions.