PFEC induced cell apoptosis and increase caspase 3 activity.

<p>B16F10 cells were double stained with Annexin V-FITC and PI and analyzed by flow cytometry following exposure to PFEC (0.4, 0.8, and 1.6 mg/ml) for 24 h. Cisplatin (20 µg/ml) was used as a positive control for apoptosis of B16F10 cells. <i>(</i><b><i>A</i></b><i>)</i> Representative flow cytometry data of cell apoptosis. <i>(</i><b><i>B</i></b><i>)</i> Summarized data of cell apoptosis. Each value was presented as mean ± SEM of three independent experiments. <i>(</i><b><i>C</i></b><i>)</i> Caspase 3 activity was increased by PF ethanol extracts. B16F10 cells were cultured and treated with PF ethanol extracts at the indicated concentrations. 24 h after treatment, cells were harvested for caspase 3 activity measurement. *<i>p</i><0.05, **<i>p</i><0.005, ***<i>p</i><0.001.</p

PFEC increased Bad, caspase8 and caspase 9 expression in cells at the protein level, but decreased AKT expression.

<p>B16F10 cells were treated with PFEC as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0102673#pone-0102673-g008" target="_blank">Figure 8</a>. 24 h after treatment, total proteins were extracted from the treated cells and subjected to Western blotting for detecting the expression of Akt, Bad, caspase 8 and caspase 9. Data present one of three repeated experiments.</p

PF ethanol extracts inhibited tumor cell migration <i>in vitro</i>.

<p>B16F10 cells were plated and cultured overnight to reach ∼80% confluency. Cells were scratched with a 200 µl of pipette tip and then washed with culture medium. Fresh culture media containing a variety of PFEC were added to the cells. Cells were allowed to grow for an additional 24 h. The cell migration distances were imaged under a phase contrast microscope (x 40). <i>(</i><b><i>A</i></b><i>)</i> Representative images of cell migration. <i>(</i><b><i>B</i></b><i>)</i> Relative cell migration rate. The distance of cell migration overnight was measured. The relative migration rate was calculated as compared with the control group. Data is a summary of three independent of experiments. *<i>p</i><0.05.</p

PF ethanol extracts reduced the mitochondrial membrane potential of B16F10 cells.

<p>B16F10 cells were treated with DiOC6 and the intensity of DiOC6 was measured by flow cytometry. Data were representative of three independent experiments.</p

Toxicity of PFEC on human gastric cancer cell BGC-823 and human immortal gastric cells GES-1.

<p><i>(</i><b><i>A</i></b><i>)</i> Cell viability of BGC-823. Cells were treated as in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0102673#pone-0102673-g001" target="_blank">Figure 1</a>. 24 h after treatment, cell viability was detected by MTT assay. The relative cell viability was calculated using control (culture with complete medium only) as a normalizer. <i>(</i><b><i>B</i></b><i>)</i> Cell proliferation. BGC-823 cells were cultured and treated with PFEC or Cisplatin. Cell proliferation at 24, 48 and 72 hours was detected as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0102673#pone-0102673-g001" target="_blank">Figure 1</a>. <i>(</i><b><i>C</i></b><i>)</i> Cell viability of GES-1 cells. Immortalized human gastric epithelial mucosa GES-1 cells were cultured and treated with PFEC, cell viability was measured by MTT assays. <i>(</i><b><i>D</i></b><i>)</i> Cell proliferation curve of GES-1 cells. GES-1 cells were cultured and treated with PFEC, cell viability was detected using MTT assays. Data shown is a summary of three independent experiments. **<i>p</i><0.005 and ***<i>p</i><0.001.</p

PF suppressed melanoma growth <i>in vivo</i>.

<p><i>(</i><b><i>A</i></b><i>)</i> Body weight of mice. C57BL/6 mice (n = 10/group) were inoculated with B16F10 melanoma cells. On the same day, mice were treated with PBS, PFEC, or Cisplatin. Body weight of mice was measured every three days using a scale. <i>(</i><b><i>B</i></b><i>)</i> Tumor growth curve. Tumor size was measured every three days with a caliper. <i>(</i><b><i>C</i></b><i>)</i> Tumor weight. At the end point of the experiments, tumor tissues were collected and weighed. ***<i>p</i><0.001.</p

PFEC regulated gene expression <i>in vitro.</i>

<p>B16F10 cells were plated and treated with PFEC. 24-PCR. Gene expression was calculated using ΔΔCt and normalized with the RPMI 1640 treated group. GAPDH was used as an internal control. <i>(</i><b><i>A</i></b><i>)</i> Akt; <i>(</i><b><i>B</i></b><i>)</i> PI3K; <i>(</i><b><i>C</i></b><i>)</i> Bcl-2; <i>(</i><b><i>D</i></b><i>)</i> Bcl-xL; <i>(</i><b><i>E</i></b><i>)</i> Bax; <i>(</i><b><i>F</i></b><i>)</i> Bid, (<i>G</i>) Cyclin D1, and (<i>H</i>) PCNA. Experiments were repeated three times independently. *<i>p</i><0.05, **<i>p</i><0.005, ***<i>p</i><0.001.</p

18 h cold ischemia-reperfusion caused I/R injury in heart grafts.

<p>Donor hearts were isolated from C57BL/6 mice, infused with UW solution and preserved in UW solution at 4°C for 18 h. After 18 h preservation, donor hearts were implanted into syngeneic recipient C57BL/6 mice. At day 2 after transplantation, the heart grafts were harvested and fixed in 10% formalin. The paraffin heart sections were subjected to HE staining, TUNEL assay, and MPO assay. (<b>A</b>) HE staining. (<b>B</b>) TUNEL assay for apoptosis. (<b>C</b>) MPO assay. Representative images were from experiments.</p

Paternal and Maternal Genetic Analysis of a Desert Keriyan Population: Keriyans Are Not the Descendants of Guge Tibetans

<div><p>The Keriyan people live in an isolated village in the Taklimakan Desert in Xinjiang, Western China. The origin and migration of the Keriyans remains unclear. We studied paternal and maternal genetic variance through typing Y-STR loci and sequencing the complete control region of the mtDNA and compared them with other adjacent populations. Data show that the Keriyan have relatively low genetic diversity on both the paternal and maternal lineages and possess both European and Asian specific haplogroups, indicating Keriyan is an admixture population of West and East. There is a gender-bias in the extent of contribution from Europe vs. Asia to the Keriyan gene pool. Keriyans have more genetic affinity to Uyghurs than to Tibetans. The Keriyan are not the descendants of the Guge Tibetans.</p></div

Median Joining network of Keriyans and other 14 referenced populations using 13 Y-STR haplotypes.

<p>A Median Joining network was constructed using the same 13 Y-STR loci as <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0100479#pone-0100479-g002" target="_blank">Figure 2</a>. Node size is proportional to frequencies of haplotype. Populations were labelled by different colors.</p