Synthesis and Insecticidal Activity of Enzyme-Triggered Functionalized Hollow Mesoporous Silica for Controlled Release

In the present study, enzymatic responsive controlled release formulations (CRFs) were fabricated. The CRFs were achieved by anchoring mechanically interlocked molecules using α-cyclodextrin onto the surface pore rims of hollow mesoporous silica (HMS). The CRFs were characterized using Fourier transform infrared spectroscopy, scanning electron microscopy, transmission electron microscopy, and thermogravimetric analysis. The results showed that the CRFs had extraordinary loading ability for chlorantraniliprole (42% w/w) and could effectively preserve chlorantraniliprole against degradation under thermal conditions and UV radiation. The CRFs have been proven to be enzyme-sensitive. The release ratio of chlorantraniliprole from CRFs can be accelerated observably when external α-amylase was introduced. The persistence of CRFs was evaluated by regular sampling feeding experiment using <i>Plutella xylostella</i> as the target insect. The results showed that the larval mortality of <i>P. xylostella</i> was much higher than that of Coragen under all concentrations after 14 days, which proved that CRFs had remarkable persistence

Aurora-A induced PAK7 expression in esophageal cancer cells.

<p>A: The heatmap of the array results. Left: 293TREX-Aur(DOX+/−); Right: EC9706-Aur/P4. B: Genes with upregulation or downregulation of more than 2-fold in both 293TREX-Aur and EC9706-Aur cells compared with control cells. The negative inverse results are reported as fold-downregulation. C. Aurora-A induced PAK7 mRNA expression. D. Aurora-A elevated PAK7 protein level. E. EC9706 cells were treated with 50 nM MLN8237 for 24 hrs and harvested for detection of PAK7 expression by Real-time PCR. F. Immunohistochemical analysis of Aurora-A and PAK7 expression in esophageal cancer samples. Representative cases of weak and strong staining for Aurora-A and PAK7 are shown. The original magnification: ×100 (pictures), and ×400 (insets).</p

Knockdown of PAK7 increased cell apoptosis in Aurora-A-overexpressing cancer cells.

<p>A. EC9706-AURKA cells were treated with 80 nM PAK7 or Negative Control (NC) siRNA for 48 h and then exposed to 5 µM CDDP for 36 h. The cells were harvested and stained with AnnexinV/PI to determine apoptosis rates using flow cytometer. The efficiency of PAK7 knockdown was determined by Western blotting. B. TUNEL assay of EC9706 cells with different treatment. C. Percentage of TUNEL positive cells. Data represented the mean percentage from three independent experiments.</p

Aurora-A induced PAK7 expression through E2F1 regulation.

<p>A. Real-time PCR analysis of PAK7 mRNA expression in EC9706 cells transfected with E2F1 siRNA. B. Chromatins prepared from 293TREX-Aur cells treated with or without DOX were subjected to immunoprecipitation with either an anti-E2F1 antibody or mouse IgG. The precipitated DNA was detected by PCR assay. Input DNA of the two samples were subjected to PCR to ensure that equal amounts of total DNA were used in the immunoprecipitation.</p

Summary of Aurora-A and PAK7 expression in human esophageal cancers.

<p>The levels of Aurora-A and PAK7 expression were determined in 121 surgical specimens of primary esophageal cancers. The correlation was analyzed using a Spearman's rank correlation test, p<0.001.</p><p>Summary of Aurora-A and PAK7 expression in human esophageal cancers.</p

Aurora-A overexpression enhanced anti-apoptotic effects of CDDP in EC9706 cells.

<p>A: EC9706-P4 and EC9706-Aur cells were plated at a density of 5000 cells/well into 96-well plates and then subjected to CDDP treatment at 10 µM for 55 hours. The cell viablity was analyzed by MTT assay. B: EC9706-P4 and EC9706-Aur cells were treated with CDDP at 5 µM for 36 hours. Cells were harvested and stained for Annexin V/PI and subjected to flow cytometer for determination of apoptosis rates. C. EC9706, EC9706-Aurk and EC9706-Aurk-KD were treated with 10 µM CDDP for 48 hrs. Cells were harvested and fixed with 70% ethol for 1 hr, then cells were stained with PI and analyzed with Flow cell cytometer. D. EC9706-Aurk and EC9706-Aurk-KD cells were treated with different doses of CDDP and MTT assay was performed to determine the viability of the cells.</p

Curcumin down-regulates DNMT1 mRNA and protein expression in myeloid leukemia cells <i>in vitro</i> and <i>in vivo</i>.

<p>(A–D) DNMT1 and GAPDH antibodies were used for immunoblotting of total cell lysates from K562 (A), THP-1 (B), Kasumi-1 (B and C), and MV4–11 cells (B), or primary AML cells (D). These cells were either left untreated, treated with vehicle (DMSO), or treated with 10 µM curcumin (“Cur”) or 20 µM curcumin. Treatment lasted for 24 h (A, B), 7 and 24 h (C), or 72 h (D). Data shown in (A–D) are from one representative experiment of three total experiments, each with similar data. Numbers beneath each lane represent densitometric quantification of DNMT1, normalized to GAPDH. Values, depicted as percent change, were calculated as change relative to incubation with carrier control. (E, F) MV4–11 cells (E) or primary AML cells (F) were incubated with carrier containing either curcumin (curcumin was used at the concentrations indicated for MV4–11 cells, and 10 µM for primary cells) or decitabine (2.5 µM; positive control). DNMT1 transcript was assessed by RT PCR, and normalized to GAPDH internal control. *, <i>p</i><0.05; **, <i>p</i><0.01. (G) Peripheral blood mononuclear cells (PBMCs) were treated with the indicated concentrations of curcumin for 24 h. Total cell lysates were then used for immunoblot assay to detect DNMT1 and β-actin. Results from one of three donors with similar data are shown. Numbers beneath each lane represent densitometric quantification of DNMT1, normalized to β-actin.</p

Curcumin inhibits p65 and Sp1 protein expression and the association of Sp1 with the <i>DNMT1</i> promoter.

<p>(<b>A</b>) After MV4–11 cells were treated with curcumin (10 µM) for the indicated periods of time, cytoplasmic and nuclear extracts were analyzed for p65 protein expression via immunoblotting. Numbers beneath each lane represent densitometric quantification of p65 protein expression, following normalization to β-actin (cytoplasmic) or H2B (nuclear) expression. (B) Whole cell lysates were prepared from MV4–11 cells incubated for 24 h with carrier alone or carrier containing curcumin (concentrations as indicated). Sp1 protein expression was assessed via immunoblotting, quantified via densitometry, and normalized to β-actin loading control. Data shown in (A) and (B) are from one representative experiment of three total experiments, each with similar data. (C) A ³²P-labeled probe, containing two Sp1 binding sites on the <i>DNMT1</i> promoter (top panel) and an Sp1 antibody were used to perform EMSA assays using nuclear extracts from MV4–11 cells, either before treatment (bottom, left panel) or following incubation for 72 h or 48 h (not shown) with carrier alone or carrier containing the indicated concentration of curcumin (bottom, right panel).</p

The anti-tumor effect of curcumin on MV4–11 tumor cells engrafted in nude mice.

<p>(A–D) MV4–11 cells were engrafted in nu/nu mice. Mice were then treated daily (5 days/week), with IP doses of either 100 mg/kg curcumin formulated in vehicle (DMSO, ethanol and PBS) or formulation vehicle alone (6 mice per group). (A, B) Tumor sizes were measured (as indicated in Materials and Methods) and recorded on days 1, 8, 15, 22 and 29. (A) Tumor growth in each mouse as it occurred over time is depicted here as individual dots (empty circles represent individual mice treated with carrier control, and solid dots represent individual curcumin-treated mice). Trend lines represent mean tumor size of all mice from the control group (grey) or the curcumin-treated group (black). (B) The mice described above were sacrificed on day 29 to isolate tumors. Tumors were weighed, and the average percent reduction in tumor weight observed in the curcumin-treated group was calculated relative to the average tumor weight of the mice from the vehicle control group. *, <i>p</i><0.05. (C, D) MV4–11-engrafted tumor tissues from the mice described above were excised 24 h after administration of the last dose of either curcumin or vehicle alone to measure DNMT1 protein expression. DNMT1 protein was measured via immunoblotting (C). Data for three individual representative mice from each treatment group are shown. Numbers beneath each lane represent quantification of DNMT1 by densitometry, normalized to β-actin. (D) <i>DNMT1</i> mRNA expression was assessed via real-time RT-PCR and normalized to <i>GAPDH</i> internal control. Values depicted as the percent change were calculated as the change relative to treatment with carrier control. **, <i>p</i><0.01.</p

Curcumin increases the proportion of cells in SubG1 and G1 phases of the cell cycle, induces caspase cleavage, and inhibits AML cell growth <i>in vitro</i>.

<p>(A, B) MV4–11 cells were placed in serum-free medium for 48 h to synchronize cell cycle. Cells were then treated with carrier (control) or curcumin (10 µM) for 48 h, and cell cycle distribution was determined by flow cytometry. The percentages of cells in SubG1, G1, S, G2/M phases of the cell cycle were calculated for each treatment condition. Gating for each phase was performed as shown in histograms from a representative experiment in (A). Average percentages of cells in subG1 and G1 (after excluding subG1) from three independent experiments are summarized in (B). (C) MV4–11 cells were treated with curcumin for time periods as indicated. Total cell lysates were used for immunoblot assay with an antibody against cleaved caspase-3, cleaved caspase-9, or β-actin (for normalization). Results from one of three experiments with similar data are shown. Numbers beneath each lane represent densitometric quantification of cleaved caspase-3 or -9, normalized to β-actin. D) MV4–11 cells were treated with carrier alone (DMSO) or carrier containing curcumin at the indicated concentrations. After 24 h, 48 h, or 72 h, proliferation was assessed using CellTiter 96 Aqueous One Solution Reagent, and is presented here as absorbance measured by a spectrophotometer. *, <i>p</i><0.05, **, <i>p</i><0.01.</p