Journal: Journal of Cellular and Molecular Medicine

Article Title: Mesenchymal stem cells expressing TRAIL lead to tumour growth inhibition in an experimental lung cancer model

doi: 10.1111/j.1582-4934.2008.00317.x

Figure Lengend Snippet: MSCs transduced with Ad.TR can induce apoptosis in A549 cells. ( A ) MSCs were transduced with Ad.BGal (100 pfu/cell) or Ad.TR (100 pfu/cell). After 48 hrs MSCs were trypsinized and 10 4 of these cells were added to wells that had been seeded the day before with A549 cells at 10 5 cells per well. After 48 hrs, wells were trypsinized and the complete mixed cell population was measured by Nicoletti apoptosis assay. A549 cells and MSCs cultured alone as well as A549 cells mixed with MSCs transduced with Ad.BGal showed only background apoptosis of below 5%. In contrast, in A549 mixed with MSCs transduced with Ad.TR apoptosis rates of almost 30% could be measured. This apoptosis could be inhibited by TRAIL neutralizing antibodies (α-TR-ab) and the pan-caspase inhibitor zVAD. Numbers represent mean values of three samples ± standard deviation. ** P < 0.001. ( B ) MSCs cocultured with A549 cells represent approximately 1% of the final cell population. 10 4 untransduced or Ad.BGal transduced (100 pfu/cell) MSC cells were added to wells that were unseeded or had been seeded the previous day with 10 5 A549 cells. After 48 hrs, cells were detached from wells using trypsin free buffer (Specialty Media, NJ, USA). Cells were stained with anti-CD105 and analysed by flow cytometry. The isotype control for a 1:10 MSC:A549 mix (MSC+A549 ISO) was, as expected, negative for CD105, whereas 98% of all cells in a pure MSC culture (MSC) were CD105 positive. A549 cells only (A549) showed a weak CD105 signal in 0.5% of all cells. In 1:10 mixes with MSCs (MSC+A549) or MSCs transduced with Ad.BGal (A549+MSCβGal) the CD105 signal increased to about 1.5% 48 hrs after mixing and coculturing. Please note that graph is depicted in log-scale. ( C ) The experiment was performed as described above , however, untransduced and transduced (Ad.BGal and Ad.TR) MSC cells were added at 10 4 (1:10) (black bars) and 3.3 × 10 3 (1:30) (white bars) cells per well. After 48 hrs, cells were analysed by Nicoletti apoptosis assay proving that even at a lower ratio against the target cancer cells, MSCs expressing TRAIL can trigger significant levels of apoptosis. Numbers represent mean values of 12 samples ± standard deviation. ** P < 0.001. ( D ) MSCs were transduced with Ad.EGFP (100 pfu/cell) (black bars) or Ad.TR (100 pfu/cell) (white bars). After 48 hrs, MSCs were harvested and 10 4 of these cells were mixed with 10 5 pre-seeded primary human fibroblasts (Fib). After 48 hrs, apoptosis was determined by Nicoletti apoptosis assay. HCT116 cells (HCT) served as positive controls in this experiment.

Article Snippet: The pan-caspase inhibitor zVAD was bought from Santa Cruz (Santa Cruz, CA, USA).

Techniques: Transduction, Apoptosis Assay, Cell Culture, Standard Deviation, Staining, Flow Cytometry, Control, Expressing

Journal: Molecular Cancer

Article Title: Circular RNA circNHSL1 promotes gastric cancer progression through the miR-1306-3p/SIX1/vimentin axis

doi: 10.1186/s12943-019-1054-7

Figure Lengend Snippet: CircNHSL1 promotes gastric cancer progression by serving as a miRNA sponge of miR-1306-3p. a Pearson correlation analysis determined the significantly negative correlation between the levels of circNHSL1 and miR-1306-3p in 61 paired gastric cancer tissues (p < 0.01). b The effects of circNHSL1 on the expression of SIX1 was detected by qRT-PCR. c The effects of mimics, inhibitor and negative control on the luciferase activities were detected in gastric cancer cells after transfecting luciferase reporter plasmids with wild type of WT, Mutant or negative control. d Schematic illustration of the sequence of wild type of SIX1 3′-UTR (WT) and mutant sequences on the complementary sites of SIX1 3′-UTR with miR-1306-3p (Mutant). e and f Anti-AGO2 RIP assay was performed in MKN-28 cells after transfection with mimics or negative control, followed by agarose gel electrophoresis ( e ) and qRT-PCR ( f ) to detect the expression of circNHSL1 and miR-1306-3p. g The effects of circNHSL1 and miR-1306-3p on the luciferase activities of wild type of SIX1 mRNA 3′-UTR in gastric cancer cells were detected. h and i The effects of circNHSL1 and miR-1306-3p on the mRNA and protein expressions of SIX1 and Vimentin were detected by qRT-PCR and western blotting. j and k The effects of circNHSL1 and miR-1306-3p on the mobility, migration and invasion were confirmed by wound healing and transwell assays. All data are presented as the mean ± SEM of three experiments. ** p < 0.01

Article Snippet: The mimics, inhibitor and negative controls for hsa-miR-1306-3p were purchased from RiBoBio (Guangzhou, China).

Techniques: Expressing, Quantitative RT-PCR, Negative Control, Luciferase, Mutagenesis, Sequencing, Transfection, Agarose Gel Electrophoresis, Western Blot, Migration

Journal: Molecular Cancer

Article Title: Circular RNA circNHSL1 promotes gastric cancer progression through the miR-1306-3p/SIX1/vimentin axis

doi: 10.1186/s12943-019-1054-7

Figure Lengend Snippet: MiR-1306-3p is down-regulated in gastric cancer tissues and correlated with the progression and poor prognosis. The expression of miR-1306-3p was detected by ISH with TMA including 54 paired gastric cancer tissues and paired normal tissues. a Represent images of miR-1306-3p expression in normal gastric mucosa and gastric cancer tissues with Stage II, Stage III, M0, M1, G1, G2, G3, pT2, pT3 and pT4. b The level of miR-1306-3p in gastric cancer tissues was significantly lower than in normal gastric tissues. c-f The level of miR-1306-3p in gastric cancer tissues with Stage II ( c ), M0 ( d ), well differentiation ( e ) and pT2 and pT3 ( f ) was significantly higher than these with Stage III, M1, moderate and poor differentiation and pT4. g - h Kaplan-Meier survival analysis (log-rank test) showed that gastric cancer patients with low miR-1306-3p expression have a lower OS and DFS than these with high miR-1306-3p expression ( p < 0.01). All data are presented as the mean ± SEM. ** p < 0.01

Article Snippet: The mimics, inhibitor and negative controls for hsa-miR-1306-3p were purchased from RiBoBio (Guangzhou, China).

Techniques: Expressing