The Inhibitory Effect of Pseudolaric Acid B on Gastric Cancer and Multidrug Resistance via Cox-2/PKC-α/P-gp Pathway

<div><p>Aim</p><p>To investigate the inhibitory effect of pseudolaric acid B on subcutaneous xenografts of human gastric adenocarcinoma and the underlying molecular mechanisms involved in its multidrug resistance.</p><p>Methods</p><p>Human gastric adenocarcinoma SGC7901 cells and drug-resistant SGC7901/ADR cells were injected into nude mice to establish a subcutaneous xenograft model. The effects of pseudolaric acid B with or without adriamycin treatment were compared by determining the tumor size and weight. Cyclo-oxygenase-2, protein kinaseC-α and P-glycoprotein expression levels were determined by immunohistochemistry and western blot.</p><p>Results</p><p>Pseudolaric acid B significantly suppressed the tumor growth induced by SGC7901 cells and SGC7901/ADR cells. The combination of pseudolaric acid B and the traditional chemotherapy drug adriamycin exhibited more potent inhibitory effects on the growth of gastric cancer in vivo than treatment with either pseudolaric acid B or adriamycin alone. Protein expression levels of cyclo-oxygenase-2, protein kinaseC-α and P-glycoprotein were inhibited by pseudolaric acid B alone or in combination with adriamycin in SGC7901/ADR cell xenografts.</p><p>Conclusion</p><p>Pseudolaric acid B has a significant inhibitory effect and an additive inhibitory effect in combination with adriamycin on the growth of gastric cancer in vivo, which reverses the multidrug resistance of gastric neoplasm to chemotherapy drugs by downregulating the Cox-2/PKC-α/P-gp/mdr1 signaling pathway.</p></div

Cox-2, PKC-α, and P-gp expression levels of xenografts in different groups (×400).

<p>A: expression of Cox-2 in mice xenografts B: expression of PKC-α in mice xenografts C: expression of P-gp in mice xenografts D: negative control E: xenograft by HE F: normal gastric tissue by HE. Tissues were fixed, embedded, mounted and stained by the SP method or HE staining using standard procedures. Scale bar, 25 µm.</p

Average optical density of Cox-2, PKC-α and P-gp in different treatment groups (mean ± SD).

<p>Notes: * p<0.05 vs control group.</p><p>n: number of animals.</p><p>PAB, Pseudolaric acid B; ADR, adriamycin; SD, standard deviation.</p><p>Average optical density of Cox-2, PKC-α and P-gp in different treatment groups (mean ± SD).</p

Inhibitory effects of PAB on expressions of Cox-2, PKC-α and P-gp in tumor tissues.

<p>A: representative immunoblots of Cox-2, PKC-α, P-gp compared with β-actin in different treatment groups. (lane 1): NS control group (SGC7901/ADR) (lane 2): TWEEN control group (SGC7901/ADR) (lane 3): ADR group (SGC7901/ADR) (lane 4): PAB group (SGC7901/ADR) (lane 5): PAB+ADR group (SGC7901/ADR) (lane 6): NS control group (SGC7901). B: relative expression levels of Cox-2 (B1), PKC-α (B2), and P-gp (B3) in different treatment groups. The tumors were isolated and homogenized to measure the protein level, and β-actin was used as an internal control. Each immunoblot was representative of three distinct experiments with similar results. *<i>p</i><0.05 vs control group.</p

Antitumor effects of PAB and/or ADR on nude mice xenografts of SGC7901 cells and SGC7901/ADR cells.

<p>Notes: * p<0.05 vs control group.</p><p>Rtv/Ctv (%)  =  (Relative tumor volume in treatment group/Relative tumor volume in NS control group) ×100.</p><p>Inhibition rate (%)  =  [(1-mean RTV of drug-treated group/mean RTV of NS control group) ×100].</p><p>Body weight (BW) change (%)  =  [(BW on day 20- tumor weight) - (BW on day 1)]/(BW on day 1) ×100.</p><p>PAB, Pseudolaric acid B; ADR, adriamycin.</p><p>Antitumor effects of PAB and/or ADR on nude mice xenografts of SGC7901 cells and SGC7901/ADR cells.</p

Inhibition of gastric tumor growth by pseudolaric acid B and mean body weight change of nude mice.

<p>A: growth curve of SGC7901 cell xenografts B: growth curve of SGC7901/ADR cell xenografts C: relative body weight change of mice with SGC7901 cell xenografts D: relative body weight change of mice with SGC7901/ADR cell xenografts. SGC7901 cells and SGC7901/ADR cells (2.5×10⁶/ml) were injected subcutaneously into the axillary areas of nude mice, and when evident tumors were observed, the mice received a daily dose of 25 mg/kg pseudolaric acid B and/or 1.25 mg/kg adriamycin or normal saline (control)/tween solution (control) (n = 5 per group). Tumor volumes were monitored at different time points (Day1, 3, 5, 7, 9, 11, 13, 15, 17, 19). The IR of tumor growth in the drug-treated groups were compared with that in the control groups at the end of the experiment by one-way ANOVA. *<i>p</i><0.05 vs control group.</p

Chemical structure of pseudolaric acid B (C₂₃H₂₈O₈, MW = 432.5).

<p>Chemical structure of pseudolaric acid B (C₂₃H₂₈O₈, MW = 432.5).</p

Subcutaneous xenografts of nude mice in different treatment groups.

<p>A: isolated tumors of SGC7901 cells (above) and SGC7901/ADR cells (below) B: subcutaneous xenografts of SGC7901 cell line in nude mice C: subcutaneous xenografts of SGC7901/ADR cell line in nude mice. (group 1): NS control group (group 2): TWEEN control group (group 3): ADR group (group 4): PAB group (group 5): PAB+ADR group.</p

Tau phosphorylation by GSK-3β promotes tangle-like filament morphology-2

<p><b>Copyright information:</b></p><p>Taken from "Tau phosphorylation by GSK-3β promotes tangle-like filament morphology"</p><p>http://www.molecularneurodegeneration.com/content/2/1/12</p><p>Molecular Neurodegeneration 2007;2():12-12.</p><p>Published online 28 Jun 2007</p><p>PMCID:PMC1936422.</p><p></p>phorylated tau (filled symbols) showed that phosphorylation altered polymerization kinetics, particularly in the first 30 minutes of the reaction. Panel A shows kinetics over the entire 20-h reaction time. Panel B shows only the first 30 minutes (the section boxed by a dotted line in panel A). Two ratios of ARA inducer:tau protein were compared: a suboptimal ratio (labeled 25 μM, representing the concentration of ARA, diamonds) and an optimal ratio (labeled 75 μM, circles). The tau protein concentration was 2 μM for all kinetic reactions. Changes in ThS fluorescence intensity (y axis) was used to indicate the extent of polymerization and measurements were in arbitrary units (a.u.). Error bars are +/- SEM. Every 40th data point was plotted for ease in interpretation

Tau phosphorylation by GSK-3β promotes tangle-like filament morphology-4

<p><b>Copyright information:</b></p><p>Taken from "Tau phosphorylation by GSK-3β promotes tangle-like filament morphology"</p><p>http://www.molecularneurodegeneration.com/content/2/1/12</p><p>Molecular Neurodegeneration 2007;2():12-12.</p><p>Published online 28 Jun 2007</p><p>PMCID:PMC1936422.</p><p></p> either a low (25 μM), or optimal (75 μM) concentration of ARA (Panels A and B, respectively). Panels C-F show polymerization reaction products in which mock-phosphorylated tau had been polymerized. In panel C and D, GSK-3β had been omitted from the phosphorylation reaction (- GSK + ATP) and in panel E and F, ATP had been omitted from the phosphorylation reaction (+ GSK – ATP). The 25 μM ARA reactions were diluted five fold, the 75 μM ARA reactions were diluted ten fold prior to grid preparation