Compound Bieshe Kang’ai inhibits proliferation and induces apoptosis in HCT116 human colorectal cancer cells

Purpose: To study the effect of Compound Bieshe Kang’ai (CBK) on proliferation and apoptosis in colorectal cancer cells.Methods: HCT116 colorectal cancer cells and FHs 74 Int intestinal cells were treated with CBK, followed by determination of cell proliferation with 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Caspase-9 and caspase-3 activities as well as protein expressions of Bcl-2 and BAX, and mRNA levels of caspase-9, caspase-3, Bcl-2 and BAX in HCT116 cells were evaluated, followed by examination of the morphological alterations of HCT116 cells with Hoechst 33342 staining.Results: CBK suppressed proliferation of HCT116 cells in a concentration- and time-dependent pattern, without cytotoxicity to FHs 74 Int cells. CBK also elevated caspase-9 and caspase-3 activities, mitigated protein translation of Bcl-2 and augmented that of BAX. It also enhanced mRNA transcriptions of caspase-9, caspase-3 and BAX, but decreased that of Bcl-2 in HCT116 cells in a  concentrationdependent manner, as well as induced cancer cell shrinkage, nuclear fragmentation and chromatin condensation.Conclusion: The findings highlight CBK as a promising therapeutic agent for colorectal cancers, by retarding proliferation and inducing apoptosis in cancer cells.Keywords: Apoptosis, BAX, Bcl-2, Cancer, Caspase, Compound Bieshe Kang’ai, Chromatin condensation, Nuclear fragmentatio

Theoretical and Experimental Studies of a PDMS Pneumatic Microactuator for Microfluidic Systems

The compact, simple, and fast-reaction pneumatic microactuator is significant for the integration and high efficiency of pneumatic systems. In this work, the structure, working principle, and multiphysical model of an on-chip pneumatic microactuator are presented. The on-chip pneumatic microactuator is mainly composed of two parts: a polydimethylsiloxane (PDMS) thin membrane and an actuated chamber. The air pressure in the actuated chamber drives the thin elastic membrane to deformation. Dynamic response mathematical models of the actuated chamber for charging and exhaust with variable volume are established, and the deformation characteristics of the polydimethylsiloxane (PDMS) actuated membrane, the capacity of the actuated chamber, and the valve opening of the on-off membrane microvalve are simulated and analyzed to explore the response characteristics of the proposed pneumatic microactuator. Samples valving analysis of the on-chip membrane microvalve and mixing performance of the micromixer integrated with the pneumatic microactuator are tested to evaluate the driving capability of the pneumatic microactuator, and the results show that the response performance of the actuated time fully satisfies the needs of a pneumatic microfluidic chip for most applications

Theoretical and Experimental Studies of a PDMS Pneumatic Microactuator for Microfluidic Systems

The compact, simple, and fast-reaction pneumatic microactuator is significant for the integration and high efficiency of pneumatic systems. In this work, the structure, working principle, and multiphysical model of an on-chip pneumatic microactuator are presented. The on-chip pneumatic microactuator is mainly composed of two parts: a polydimethylsiloxane (PDMS) thin membrane and an actuated chamber. The air pressure in the actuated chamber drives the thin elastic membrane to deformation. Dynamic response mathematical models of the actuated chamber for charging and exhaust with variable volume are established, and the deformation characteristics of the polydimethylsiloxane (PDMS) actuated membrane, the capacity of the actuated chamber, and the valve opening of the on-off membrane microvalve are simulated and analyzed to explore the response characteristics of the proposed pneumatic microactuator. Samples valving analysis of the on-chip membrane microvalve and mixing performance of the micromixer integrated with the pneumatic microactuator are tested to evaluate the driving capability of the pneumatic microactuator, and the results show that the response performance of the actuated time fully satisfies the needs of a pneumatic microfluidic chip for most applications

ZHX1 Promotes the Proliferation, Migration and Invasion of Cholangiocarcinoma Cells

<div><p>Zinc-fingers and homeoboxes 1 (ZHX1) is a transcription repressor that has been associated with the progressions of hepatocellular carcinoma, gastric cancer, and breast cancer. However, the functional roles of ZHX1 in cholangiocarcinoma (CCA) have not been determined. We investigated the expression and roles of ZHX1 during the proliferation, migration, and invasion of CCA cells. <i>In silico</i> analysis and immunohistochemical studies showed amplification and overexpression of ZHX1 in CCA tissues. Furthermore, ZHX1 knockdown using specific siRNAs decreased CCA cell proliferation, migration, and invasion, whereas ZHX1 overexpression promoted all three characteristics. In addition, results suggested EGR1 might partially mediate the effect of ZHX1 on the proliferation of CCA cells. Taken together, these results show ZHX1 promotes CCA cell proliferation, migration, and invasion, and present ZHX1 as a potential target for the treatment of CCA.</p></div

ZHX1 promoted the migration of cholangiocarcinoma cells.

<p>Migration was examined using a Boyden chamber assay and a wound healing assay. (A) ZHX1 siRNA significantly inhibited the FBS-induced migrations of SNU478 and SNU1196 cells compared with SCR siRNA. Scale bar represents 50 μm. (B) The number of migrated cells were counted and SCR siRNA-treated controls were used as controls to calculate the percentage of cell migration in ZHX1-knockdown cells. The values are shown as the bar graph. Results are the means ± SEs of three independent experiments. (C) Overexpression of ZHX1 significantly increased the migration of HuCCT1 cells versus Mock cells. Scale bar represents 50 μm. (D) The number of migrated cells were counted and Mock cells were used as controls to calculate the percentage of cell migration in ZHX1-overexpressing cells. Results are the means ± SEs of three independent experiments. (E) Migration of cholangiocarcinoma cells was examined using a wound healing assay. Cells were scratched one day after seeding, washed twice with 1X PBS, and then the fresh media containing 100 ng/ml of mitomycin C and 0.1% FBS was added. Pictures were taken at 0 hour and 21 hours after scratching. Five separate experiments were performed. (F) Migration was quantified by measuring the scratch widths. Mock cells were used as controls. The bar graphs show the means ± SEs of five independent experiments. *, P < 0.05, **, P<0.01, versus SCR or Mock.</p

ZHX1 accelerated the invasion of cholangiocarcinoma cells.

<p>A matrigel invasion assay was used to examine the invasive ability of CCA cells. (A) ZHX1 siRNA decreased the FBS-induced invasion of SNU478 and SNU1196 cells compared with SCR siRNA. (B) The numbers of invaded cells were counted and SCR siRNA-treated cells were used as controls to calculate the percentage of cell invasion. Results are the means ± SEs of three independent experiments. (C) Overexpression of ZHX1 significantly increased the invasion of HuCCT1 cells versus Mock cells. Scale bar represents 50 μm. (D) The numbers of invaded cells were counted and Mock cells were used as controls to calculate the percentage of cell invasion. Results are the means ± SEs of three independent experiments. *, P < 0.05, **, P<0.01, versus SCR or Mock.</p

ZHX1 regulated EGR1 expression in cholangiocarcinoma cells.

<p>(a) Real-time PCR was used to assess EGR1 mRNA level changes. EGR1 mRNA levels were measured two days after SCR or ZHX1 siRNA transfection in SNU478 or SNU1196 cells and after plating Mock and ZHX1-overexpressing HuCCT1 cells. (b) Cell proliferation was observed 3 to 4 days after SCR or EGR1 siRNA transfection in SNU478, SNU1196 and HuCCT1 cells. SCR values were used as controls to calculate relative cell proliferations. Results are presented in the bar graphs as the means ± SEs of three independent experiments. *, P < 0.05, **, P<0.01, versus SCR or Mock.</p

ZHX1 promoted the migration of cholangiocarcinoma cells.

<p>Migration was examined using a Boyden chamber assay and a wound healing assay. (A) ZHX1 siRNA significantly inhibited the FBS-induced migrations of SNU478 and SNU1196 cells compared with SCR siRNA. Scale bar represents 50 μm. (B) The number of migrated cells were counted and SCR siRNA-treated controls were used as controls to calculate the percentage of cell migration in ZHX1-knockdown cells. The values are shown as the bar graph. Results are the means ± SEs of three independent experiments. (C) Overexpression of ZHX1 significantly increased the migration of HuCCT1 cells versus Mock cells. Scale bar represents 50 μm. (D) The number of migrated cells were counted and Mock cells were used as controls to calculate the percentage of cell migration in ZHX1-overexpressing cells. Results are the means ± SEs of three independent experiments. (E) Migration of cholangiocarcinoma cells was examined using a wound healing assay. Cells were scratched one day after seeding, washed twice with 1X PBS, and then the fresh media containing 100 ng/ml of mitomycin C and 0.1% FBS was added. Pictures were taken at 0 hour and 21 hours after scratching. Five separate experiments were performed. (F) Migration was quantified by measuring the scratch widths. Mock cells were used as controls. The bar graphs show the means ± SEs of five independent experiments. *, P < 0.05, **, P<0.01, versus SCR or Mock.</p

ZHX1 regulated the proliferation of CCA cells.

<p>(A) The effect of knockdown or overexpression of ZHX1 on the level of mRNA was examined by real-time PCR. For ZHX1 knockdown, CCA cells were treated with 100 nM of scrambled siRNA (SCR) or ZHX1 siRNA. SCR siRNA-treated samples were used as controls. For the gain-of-function study, ZHX1-overexpressing CCA cells (ZHX1-over) were generated from HuCCT1 cells, and empty control vector-expressing (Mock) cells were used as a control. GAPDH was used as an internal control. (B) Knockdown and overexpression efficiencies were determined by western blotting. (C) ZHX1 protein levels were quantified using image J software, and β-actin was used as an internal control. SCR siRNA-treated controls in the knockdown study, and Mock cells in the gain-of-function study were used as controls. (D) Cell proliferation was measured 3 to 5 days after SCR or ZHX1 siRNA transfection. SCR siRNA-treated controls in the knockdown study and Mock cells in the gain-of-function study were used as controls to calculate relative cell proliferation. Bar graphs show the means ± SEs of three independent experiments. *, P < 0.05, **, P<0.01, versus SCR or Mock.</p