Treatment with 30 µM DhL induces apoptosis in HeLa cells.

<p>Unsynchronized (A) and synchronized (B) HeLa cells were treated with 0, 20, or 30 µM DhL for 24 or 48 h. DNA content was assessed by flow cytometry (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0053168#pone.0053168.s002" target="_blank">Fig. S2</a>, DNA distributions). The percentages of unsynchronized and synchronized cells in the sub-G1 phase are shown. (C) Synchronized HeLa cells were treated with 0, 20, or 30 µM DhL for 24 or 48 h. Apoptotic cells were assessed by TUNEL assay. Left: representative panels with apoptotic cells indicated by arrowheads. Bar: 50 µm. Right: percentages of apoptotic cells at 24 and 48 h. (D) Unsynchronized HeLa cells were treated as in (C) and subjected to Annexin V assay. Left: representative panels with apoptotic cells stained with Annexin V (bright cells) at 24 h. Representative fields of Annexin V positive cells for 48 and 72 h treatment are shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0053168#pone.0053168.s002" target="_blank">Fig. S2</a> C. Bar: 50 µm. Right: percentages of apoptotic cells at 24, 48 and 72 h. Data represent mean ± SEM of 3 independent experiments. * p≤0.05, ** p≤0.01, *** p≤0.001 vs. control group (0 µM DhL).