Figure 6.

Activation of SARM1 by CZ-48 Induced Cell Death

(A–H) Wild-type and SARM1-overexpressing HEK-293 cells were treated by 100 μM CZ-48 for labeled time periods. (A) CZ-48 treatment induces cell blisters (black arrows) and shrinkage in cells overexpressing SARM1. (B and C) Cell viabilities were analyzed by annexin-V/PI staining combining flow cytometry (B), and the PI positive rates of all samples were plotted (C). (D) The cellular contents of cADPR (upper chart), NAD (middle chart), and ATP (lower chart) were measured by cycling assay or luminescent ATP detection assay, as described in Methods. (E and F) Mitochondrial reactive oxygen species contents were measured by MitoSOX red staining and analyzed by flow cytometry (E). The positive rates of all samples were plotted (F). (G and H) Mitochondrial membrane potential was analyzed by DIOC6(3) staining and analyzed by flow cytometry (G). The positive rates of all samples were plotted (H).

All the above experiments were repeated at least three times (means ± SDs; n = 3; Student's t test, **p < 0.01, ***p < 0.001, ****p < 0.0001).