The Effect Of Sp1 On The Invasion And Migration Of Gastrointestinal Tumor Induced By Glucose-Derived Advanced Glycation End Products

Objective: glucose-derived AGEs are reported to contribute to course of many tumors,but its underlying mechanism is still unclear now.Here,our study is to explore the effect of glucose-derived AGEs on the gastrointestinal cancer invasion and migration and the potential mechanism.Method:(1)The blood of colorectal cancer / gastric cancer patients without diabetes and normal control was collected and centrifuged,and then serum was obtained to detect the content of glucose-derived AGEs by western blot.(2)The colorectal cancer / gastric cancer tissues and paired normal tissues were collected,of which RNA and protein were extracted.RT-PCR and western blot were used to detect the expression of RAGE,Sp1,and MMP2,as well as tissue content of glucose-derived AGEs.(3)Human colorectal cancer / gastric cancer specimens were embedded with paraffin and sliced.Sections were subjected to immunofluorescence staining to detect the location and expression abundance of RAGE and Sp1.Furthermore,we analyzed the association between the RAGE expression and clinicopathological factors of colorectal cancer / gastric cancer patients.(4)8 colorectal cancer / gastric cancer cells were cultured in vitro,of which RNA and protein were extracted for detection of RAGE expression by RT-PCR and western blot.The suitable cells were chose for the subsequent experiments.(5)In vitro,colorectal cancer / gastric cancer cells were treated with different concentrations of glucose-derived AGEs and RAGE-specific antibody to observe the effect on invasion and migration of colorectal cancer / gastric cancer cells.(6)Sp1-specific si RNAs were constructed and transfected colorectal cancer / gastric cancer cells to observe the effect on metastasis induced by glucose-derived AGEs and expression of RAGE and MMP2.(7)RAGE-specific sh RNAs were used to stably know-down the RAGE expression of colorectal cancer / gastric cancer cell.The RAGE stably know-down colorectal cancer cells were injected in portal vein of nude mouse to establish the colorectal liver metastasis model.30 mg/kg glucose-derived AGEs were administrated by intraperitoneal injection every other day for two months.(8)RAGE-specific sh RNAs were used to stably know-down the RAGE expression of gastric cancer cell.Nude mouse receied an intraperitoneal injection of the RAGE stably know-down gastric cancer cells to establish the peritoneal metastasis model of gastric cancer.30 mg/kg glucose-derived AGEs were administrated by intraperitoneal injection every other day for two months.(9)Hep3B and Hep G2 were treated with the 200 ?g/ml glucose-derived AGEs to observe the changes of RAGE expression via western blot.The AGEs-sensitive cell was chose for subsequent experiment.(10)Hep3B was treated with different concentrations of glucose-derived AGEs to observe the effect on the lipid deposition by red oil O staining.(11)The cell protein of Hep3 B treated with different concentrations of glucose-derived AGEs was extracted to assay the effect of AGEs on protein expression of RAGE,Sp1,and SREBP1 c.Result:(1)Glucose-derived AGEs level in serum and tumor tissues of colorectal cancer patients,as well as expression of RAGE,Sp1,and MMP2 increased significantly.Glucose-derived AGEs treatment induced the invasion and migration of colorectal cancer cell in vitro and promoted the colorectal liver metastasis in vivo.Concomitantly,AGEs up-regulate RAGE,Sp1,and MMP2 expression.(2)There were accumulation of glucose-derived AGEs in sera and cancerous tissues,as well as high expression of RAGE,Sp1,and MMP2 in gastric cancer tissues.AGEs treatment can promote the metastasis of gastric cancer in vitro and in vivo,as well as up-regulated the expression of RAGE,Sp1,and MMP2.(3)Glucose-derived AGEs can promote the lipid deposition in Hep3 B cells and up-regulate RAGE,Sp1,and SREBP1 c expression in a dose-dependent manner.