C1GALT1 modifies O-glycans on integrin β1.

<p>(<i>a</i>) Integrin β1 carried O-glycans in HCC cells. Lysates of HA22T cells (0.5 mg) were treated with neuraminidase (Neu) and/or PNGaseF and then pulled down (PD) by PNA agarose beads. The pulled down proteins were separated by 6% SDS-PAGE and analyzed by immunoblotting (IB) with anti-integrin β1 antibody. (<i>b</i>) C1GALT1 enhanced PNA binding to integrin β1 in HCC36 cells. (<i>c</i>) Knockdown of C1GALT1 suppressed PNA binding to integrin β1 in HA22T cells. Cell lysates (1.2 mg) were immunoprecipitated (IP) by anti-integrin β1 antibody, and were treated with (+) or without (−) neuraminidase. Proteins were separated by 8% SDS-PAGE, and then blotted with PNA or anti-integrin β1 antibody. Non-specific mouse IgG was used as control.</p

C1GALT1 regulates integrin β1 activity and signaling.

<p>(<i>a</i>) The activity of integrin β1 was enhanced in C1GALT1 overexpressing cells. Mock and C1GALT1 transfectants were analyzed by flow cytometry with anti-integrin β1 antibody (left) or anti-activated integrin β1 (HUTS-21) antibody (right). Non-specific mouse IgG was used as a background fluorescence control (dash line). The mean fluorescence intensities (MFI) are shown at the bottom. Results are represented as means ± SD from three independent experiments. * <i>P</i><0.05. (<i>b</i>) Knockdown of C1GALT1 suppressed integrin β1 activity. Control (Ctr sh) and C1GALT1 knockdown (C1GALT1 sh6 and C1GALT1 sh8) cells were analyzed by flow cytometry with the indicated antibodies. The average of MFI is shown at the bottom. Results are represented as means ± SD from three independent experiments. * <i>P</i><0.05. (<i>c</i>) C1GALT1 regulated integrin β1-induced FAK activation. p-FAK (Tyr397) and total FAK in C1GALT1 overexpressing cells (left) and knockdown cells (right) were analyzed by Western blotting. Transfectants were seeded on BSA or collagen IV (Col-IV)-coated plates for 0.5 h. The integrin β1 blocking antibody (P4C10, 2 µg/ml) was incubated with cells for 10 min before seeding on collagen IV-coated plates. GAPDH was used as a loading control.</p

C1GALT1 induces HCC cell adhesion, migration and invasion through integrin β1.

<p>(<i>a</i>) The effects of the integrin β1-blocking antibody, P4C10, on C1GALT1 overexpressing cells. HCC cells were pre-treated with the indicated concentration of P4C10 or control IgG for 10 min. C1GALT1-induced cell-collagen IV adhesion (left), migration (middle), and invasion (right) were suppressed by the integrin β1-blocking antibody. Results are represented as means ± SD from three independent experiments. **<i>P</i><0.01. (<i>b</i>) The effects of the integrin β1-blocking antibody on C1GALT1 knockdown cells. P4C10 inhibited cell-collagen IV adhesion (left), migration (middle), and invasion (right) in cells transfected with control siRNA (Ctr si). Results are shown as means ± SD. * <i>P</i><0.05; **<i>P</i><0.01.</p

C1GALT1 regulates lung metastasis of HCC cells in NOD/SCID mice.

<p>(<i>a</i>) Stable overexpression and knockdown of C1GALT1 in HCC cells. The protein expression levels of C1GALT1 were analyzed by Western blotting. (<i>b</i>) Effects of C1GALT1 on lung metastasis. Metastatic tumors were increased in the C1GALT1 overexpressed group (left) and decreased in the C1GALT1 knockdown groups (right). Representative image of the excised lungs are shown at the bottom, n = 6 for each group. Results are shown as means ± SD. * <i>P</i><0.05; **<i>P</i><0.01. Blue arrows indicate the location of the tumor nodules on the lung surface. (<i>c</i>) H&E staining and immunohistochemistry of paraffin-embedded lung sections. Representative images (upper) and amplified images (middle) are shown. Immunostaining revealed the expression of C1GALT1 in metastatic tumors (bottom). The red dash line indicates the location of metastatic tumors. Scale bars  = 200 µm.</p

Cosynthesis of Cargo-Loaded Hydroxyapatite/Alginate Core–Shell Nanoparticles (HAP@Alg) as pH-Responsive Nanovehicles by a Pre-gel Method

A new core–shell nanostructure consisting of inorganic hydroxyapatite (HAP) nanoparticles as the core and organic alginate as the shell (denoted as HAP@Alg) was successfully synthesized by a pre-gel method and applied to pH-responsive drug delivery systems (DDS). HAP@Alg nanoparticles have the advantages of hydroxyapatite and alginate, where hydroxyapatite provides pH-responsive degradability, and alginate provides excellent biocompatibility and COOH functionality. Through the subsequent addition of CaCl₂ and phosphate solutions to the alginate solution, HAP@Alg nanoparticles with controllable particle sizes (ranging from 160 to 650 nm) were obtained, and their core–shell structure was confirmed through transmission electron microscopy (TEM) observation. Rhodamine 6G (R6G), a positively charged dye, was selected as a model drug for pH-sensitive DDS. R6G was encapsulated in the HAP/Alg nanoparticles upon synthesis, and its loading efficiency could reach up to approximately 63.0%. The in vitro release behavior of the loaded R6G at different pH values was systematically studied, and the results indicated that more R6G molecules were released at lower pH conditions. For example, after releasing for 8 h, the release amount of R6G at pH 2.0 was 2.53-fold the amount at pH 7.4. We attributed this pH-sensitive release behavior to the dissolution of the HAP core in acidic conditions. The results of the MTT assay and confocal laser scanning microscopy indicated that the HAP@Alg were successfully uptaken by liver cancer cells (HepG2) without apparent cytotoxicity. The synthesized HAP@Alg nanoparticles show great potential as drug nanovehicles with high biocompatibility, enhanced drug loading, and pH-responsive features for future intracellular DDS

Influence of H. pylori status on overall survival and relapse free survival.

<p>Overall survival (a), relapse free survival (b).</p

Influence of H. pylori status on overall survival according to AJCC 7th stages.

<p>stage II/III (a), stage II/IIIa/IIIb (b), stage II/IIIa(c).</p

Predictive factors for overall survival in multivariate Cox proportional-hazard analysis in patients with gastric cancer.

<p>Predictive factors for overall survival in multivariate Cox proportional-hazard analysis in patients with gastric cancer.</p

Characteristics of patients with gastric cancer assessed by <i>H</i>. <i>pylori</i> status.

<p>Characteristics of patients with gastric cancer assessed by <i>H</i>. <i>pylori</i> status.</p

Predictive factors for overall survival in multivariate Cox proportional-hazard analysis of patients with stage I, II or III gastric cancer.

<p>Predictive factors for overall survival in multivariate Cox proportional-hazard analysis of patients with stage I, II or III gastric cancer.</p