title: [Molecular mechanism of cisplatin to enhance the ability of TRAIL in reversing multidrug resistance in gastric cancer cells].
abstract: OBJECTIVE
  To study the molecular mechanism of cisplatin to enhance the ability of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) in reversing multidrug resistance in vincristine-resistant human gastric cancer SGC7901/VCR cells.
  
  
  METHODS
  MTT assay was used to measure the 50% inhibiting concentration (IC₅₀) and cell survival in SGC7901 and SGC7901/VCR cells after different treatments. SGC7901/VCR cells were treated with different concentrations of DDP, different concentrations of TRAIL alone or in combination, and then the mRNA and protein levels of several genes were determined by RT-PCR, RT-qPCR and Western-blot analysis. After targeted silencing with specific siRNA and transfection of recombinant plasmid c-myc into the SGC7901/VCR cells, the mRNA and protein levels of DR4, DR5 and c-myc were determined by RT-PCR and Western-blot analysis.
  
  
  RESULTS
  After combined treatment with TRAIL and DDP of the SGC7901/VCR cells, the IC₅₀ of VCR, DDP, ADM, and 5-Fu treatment was significantly decreased compared with the control group or TRAIL-treated group (P < 0.05). After treatment with 0, 10, 50 ng/ml TRAIL in combination with 0.4 µg/ml DDP, the SGC7901/VCR cells showed significantly higher activation of caspase 3, down-regulation of DNA-PKcs/Akt/GSK-3β signaling pathway, and higher inhibition of MDR1(P-gp) and MRP1 than those treated with TRAIL alone (P < 0.01 for all). The mRNA and protein levels of DR4, DR5, c-myc were significantly decreased after silencing c-myc with specific siRNA in the SGC7901/VCR cells (P < 0.01 for all), and were significantly increased after transfection of recombinant plasmid c-myc into the SGC7901/VCR cells (P < 0.01 foe all). After the treatment with 10 ng/ml TRAIL, 0.25 µg/ml DDP + 10 ng/ml TRAIL and 0.5 µg/ml DDP + 10 ng/ml TRAIL, the relative expression level of c-myc protein in the SGC7901/VCR cells was 0.314 ± 0.012, 0.735 ± 0.026, and 0.876 ± 0.028, respectively, and the relative expression of cytochrome C was 0.339 ± 0.036, 0.593 ± 0.020 and 0.735 ± 0.031, respectively, and the relative expression levels of DR4, DR5, active-caspase 3 and active-caspase 9 in the SGC7901/VCR cells were also increased along with increasing DDP concentrations.
  
  
  CONCLUSIONS
  The activation of DNA-PKcs/Akt/GSK-3β signaling pathway and high expression of MDR1 and MRP1 play an important role in the multi-drug resistance properties of SGC7901/VCR cells. After combining with TRAIL, DDP can enhance the expression of DR4 and DR5 through up-regulating c-myc and enhancing the activation of caspase 3 and caspase 9 by facilitating mitochondrial release of cytochrome C. It may be an important molecular mechanism of DDP-induced sensitization of TRAIL to reverse the multidrug resistancein SGC7901/VCR cells.
authors.affiliation: Department of Gastroenterology, the Affiliated Provincial Hospital of Anhui Medical University, Hefei 230001, China.
authors.forename: Xingchao
authors.initials: X
authors.lastname: Zhu
authors.affiliation: Department of Gastroenterology, the Affiliated Provincial Hospital of Anhui Medical University, Hefei 230001, China; Email: zkg@medmail.com.cn.
authors.forename: Kaiguang
authors.initials: K
authors.lastname: Zhang
authors.affiliation: Department of Gastroenterology, the Affiliated Provincial Hospital of Anhui Medical University, Hefei 230001, China.
authors.forename: Qiaomin
authors.initials: Q
authors.lastname: Wang
authors.affiliation: Department of Gastroenterology, the Affiliated Provincial Hospital of Anhui Medical University, Hefei 230001, China.
authors.forename: Si
authors.initials: S
authors.lastname: Chen
authors.affiliation: Department of Gastroenterology, the Affiliated Provincial Hospital of Anhui Medical University, Hefei 230001, China.
authors.forename: Yawen
authors.initials: Y
authors.lastname: Gou
authors.affiliation: Department of Gastroenterology, the Affiliated Provincial Hospital of Anhui Medical University, Hefei 230001, China.
authors.forename: Yufang
authors.initials: Y
authors.lastname: Cui
authors.affiliation: Department of Gastroenterology, the Affiliated Provincial Hospital of Anhui Medical University, Hefei 230001, China.
authors.forename: Qin
authors.initials: Q
authors.lastname: Li
mesh_terms: D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000378:metabolism; D000970:Antineoplastic Agents / Q000008:administration & dosage / Q000494:pharmacology*; D000971:Antineoplastic Combined Chemotherapy Protocols / Q000008:administration & dosage / Q000494:pharmacology*; D053148:Caspase 3 / Q000378:metabolism; D053453:Caspase 9 / Q000378:metabolism; D045744:Cell Line, Tumor; D002945:Cisplatin / Q000008:administration & dosage / Q000494:pharmacology*; D015536:Down-Regulation; D018432:Drug Resistance, Multiple / Q000187:drug effects*; D019008:Drug Resistance, Neoplasm / Q000187:drug effects*; D005472:Fluorouracil / Q000008:administration & dosage / Q000494:pharmacology; D005562:Formazans; D016259:Genes, myc; D038362:Glycogen Synthase Kinase 3 / Q000378:metabolism; D000071679:Glycogen Synthase Kinase 3 beta; D006801:Humans; D020128:Inhibitory Concentration 50; D027425:ATP-Binding Cassette, Sub-Family C Proteins / Q000378:metabolism; D009363:Neoplasm Proteins / Q000378:metabolism; D010957:Plasmids; D016271:Proto-Oncogene Proteins c-myc / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D034741:RNA, Small Interfering / Q000494:pharmacology; D053220:Receptors, TNF-Related Apoptosis-Inducing Ligand / Q000378:metabolism; D013274:Stomach Neoplasms / Q000188:drug therapy* / Q000473:pathology; D053221:TNF-Related Apoptosis-Inducing Ligand / Q000008:administration & dosage / Q000494:pharmacology*; D013778:Tetrazolium Salts; D014162:Transfection / Q000379:methods
chemical_list: D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D000970:Antineoplastic Agents; D053148:Caspase 3; D053453:Caspase 9; D002945:Cisplatin; D005472:Fluorouracil; D005562:Formazans; D038362:Glycogen Synthase Kinase 3; D000071679:Glycogen Synthase Kinase 3 beta; D027425:ATP-Binding Cassette, Sub-Family C Proteins; D009363:Neoplasm Proteins; D016271:Proto-Oncogene Proteins c-myc; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D053220:Receptors, TNF-Related Apoptosis-Inducing Ligand; D053221:TNF-Related Apoptosis-Inducing Ligand; D013778:Tetrazolium Salts; C489427:MYC protein, human; C105190:TNFRSF10A protein, human; C079782:MTT formazan; C000605506:GSK3B protein, human; C421109:multidrug resistance-associated protein 1