title: Reversal effects of pantoprazole on multidrug resistance in human gastric adenocarcinoma cells by down-regulating the V-ATPases/mTOR/HIF-1α/P-gp and MRP1 signaling pathway in vitro and in vivo.
abstract: To investigate reversal effects of pantoprazole (PPZ) on multidrug resistance (MDR) in human gastric adenocarcinoma cells in vivo and in vitro. Human gastric adenocarcinoma cell SGC7901 was cultured in RPMI-1640 medium supplemented with 10% fetal bovine serum and antibiotics in a humidified 5% CO(2) atmosphere at 37°C. Adriamycin (ADR)-resistant cells were cultured with addition of 0.8 µg/ml of ADR maintaining MDR phenotype. ADR was used to calculate ADR releasing index; CCK-8 Assay was performed to evaluate the cytotoxicity of anti-tumor drugs; BCECF-AM pH-sensitive fluorescent probe was used to measure intracellular pH (pHi) value of cells, whereas pH value of medium was considered as extracellular pH (pHe) value; Western blotting and immunofluorescent staining analyses were employed to determine protein expressions and intracellular distributions of vacuolar H(+) -ATPases (V-ATPases), mTOR, HIF-1α, P-glycoprotein (P-gp), and multidrug resistant protein 1 (MRP1); SGC7901 and SGC7901/ADR cells were inoculated in athymic nude mice. Thereafter, effects of ADR with or without PPZ pretreatment were compared by determining the tumor size and weight, apoptotic cells in tumor tissues were detected by TUNEL assay. At concentrations greater than 20 µg/ml, PPZ pretreatment reduced ADR releasing index and significantly enhanced intracellular ADR concentration of SGC7901 (P < 0.01). Similarly, PPZ pretreatment significantly decreased ADR releasing index of SGC7901/ADR dose-dependently (P < 0.01). PPZ pretreatment also decreased cell viabilities of SGG7901 and SGC7901/ADR dose-dependently. After 24-h PPZ pretreatment, administration of chemotherapeutic agents demonstrated maximal cytotoxic effects on SGC7901 and SGC7901/ADR cells (P < 0.05). The resistance index in PPZ pretreatment group was significantly lower than that in non-PPZ pretreatment group (3.71 vs. 14.80). PPZ at concentration >10 µg/ml significantly decreased pHi in SGC7901 and SGC7901/ADR cells and diminished or reversed transmembrane pH gradient (P < 0.05). PPZ pretreatment also significantly inhibited protein expressions of V-ATPases, mTOR, HIF-1α, P-gp, and MRP1, and alter intracellular expressions in parent and ADR-resistant cells (P < 0.05). In vivo experiments further confirmed that PPZ pretreatment could enhance anti-tumor effects of ADR on xenografted tumor of nude mice and also improve the apoptotic index in xenografted tumor tissues. PPZ pretreatment enhances the cytotoxic effects of anti-tumor drugs on SGC7901 and reverse MDR of SGC7901/ADR by downregulating the V-ATPases/mTOR/HIF-1α/P-gp and MRP1 signaling pathway.
authors.affiliation: Department of Gastroenterology, the Affiliated Drum Tower Hospital of Nanjing University, Medical School, Nanjing 210008, PR China.
authors.forename: Min
authors.initials: M
authors.lastname: Chen
authors.forename: Shu-Ling
authors.initials: SL
authors.lastname: Huang
authors.forename: Xiao-Qi
authors.initials: XQ
authors.lastname: Zhang
authors.forename: Bin
authors.initials: B
authors.lastname: Zhang
authors.forename: Hao
authors.initials: H
authors.lastname: Zhu
authors.forename: Vincent W
authors.initials: VW
authors.lastname: Yang
authors.forename: Xiao-Ping
authors.initials: XP
authors.lastname: Zou
mesh_terms: D053799:2-Pyridinylmethylsulfinylbenzimidazoles / Q000494:pharmacology*; D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000096:biosynthesis; D000230:Adenocarcinoma / Q000188:drug therapy; D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology* / Q000627:therapeutic use; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D004317:Doxorubicin / Q000494:pharmacology; D018432:Drug Resistance, Multiple; D019008:Drug Resistance, Neoplasm; D006801:Humans; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000378:metabolism; D051379:Mice; D008819:Mice, Nude; D000077402:Pantoprazole; D015398:Signal Transduction; D013274:Stomach Neoplasms / Q000188:drug therapy*; D058570:TOR Serine-Threonine Kinases / Q000378:metabolism; D059016:Tumor Microenvironment / Q000187:drug effects; D025262:Vacuolar Proton-Translocating ATPases / Q000378:metabolism
chemical_list: D053799:2-Pyridinylmethylsulfinylbenzimidazoles; D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D000970:Antineoplastic Agents; C497442:HIF1A protein, human; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D004317:Doxorubicin; D000077402:Pantoprazole; C546842:MTOR protein, human; D058570:TOR Serine-Threonine Kinases; D025262:Vacuolar Proton-Translocating ATPases