Mo1628 Sonichedgehog Signal Pathway Regulated by Sodium Butyrate Through Protein Kinase a in H. pylori Infected Gastric Cancer Cells

Abstract

Introduction/AimThe sonic Hedgehog (Shh) signaling pathway which regulates tissue regeneration and carcinogenesis during embryogenesis.Shh is initiated by the ligand of Hh binding to patched 1(Ptch) and conduted through transcription factors Gli1, Gli2 and Gli3. While Gli1 is thought to function as a transcriptional activator,phosphorylated Gli2 and Gli3 were processed by proteasome dependent cleavage. Gli3 undergoes phosphorylation at several sites by protein kinase A (PKA) followed by glycogen synthase kinase 3β (GSK 3β) and casein kinase 1 (CK1) before cleavage. It is well known that full-length Gli3 (F-Gli3) plays as a transcriptional activator and truncated Gli3 (R-Gli3) acts as a repressor. The aims of this study were 1. To show the effect of Shh signal activation in relation to cancer cell invasiveness. 2. To analyze the mechanism of sodium butyrate (NaB) affecting theShh signal pathways in H. pylori infected gastric cancer cells.Methods and material Human gastric cancer cells (AGS, MKN-45) were infected by CagA positive or negativeH. pylori strains for 6 hours andwere treated by corresponding inhibitors or NaBat 2μM for 48hr. CagA expression (wild type or mutant form) plasmids were used. Total protein lysates and RNA were collected. Ptch, Gli1, Gli2, Gli3 (F, R) were measured by RTPCR or immunoblot.Cell invasiveness wereconfirmed by invasion assay and wound healing assay. ResultsGli1 and Gli2 were overexpressed in gastric cancer cells infected by H. pylori in cagA dependent mode. Also, both increased cell invasiveness and over-expressed snail, decreased E-cadherin correlated well inH. pyloriinfectied gastric cancer cells.H. pylorishowed no significant effect on PKA activation whileNaBactivated PKA confirmed by kinase assay and immunoblot.NaB inhibited the Gli1 and Gli2 overexpression inH. pylori infected cells. NaBreversed F-Gli3/R-Gli3 protein expression regulating the Shh signal pathways in H. pylori infected gastric cancer cells.ConclusionsActivatedShh signal pathway is involved in cell invasiveness in H. pylori infected gastric cancer. NaB, a differentiating agent, affects the Shh pathways through PKA activation by post-translational and transcriptional regulation.