Generation of glyceraldehyde-derived advanced glycation end-products in pancreatic cancer cells and the potential of tumor promotion
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摘要
AIM To determine the possibility that diabetes mellitus promotes pancreatic ductal adenocarcinoma via glyceraldehyde(GA)-derived advanced glycation-end products(GA-AGEs).METHODS PANC-1,a human pancreatic cancer cell line,was treated with 1-4 mmol/L GA for 24 h. The cell viability and intracellular GA-AGEs were measured by WST-8 assay and slot blotting. Moreover,immunostaining of PANC-1 cells with an anti-GA-AGE antibody was performed. Western blotting(WB) was used to analyze the molecular weight of GA-AGEs. Heat shock proteins 90α,90β,70,27 and cleaved caspase-3 were analyzed by WB. In addition,PANC-1 cells were treated with GA-AGEs-bovine serum albumin(GA-AGEs-BSA),as a model of extracellular GA-AGEs,and proliferation of PANC-1 cells was measured.RESULTS In PANC-1 cells,GA induced the production of GA-AGEs and cell death in a dose-dependent manner. PANC-1 cell viability was approximately 40% with a 2 mmol/L GA treatment and decreased to almost 0% with a 4 mmol/L GA treatment(each significant difference was P < 0.01). Cells treated with 2 and 4 mmol/L GA produced 6.4 and 21.2 μg/mg protein of GA-AGEs,respectively(P <0.05 and P < 0.01). The dose-dependent production of some high-molecular-weight(HMW) complexes of HSP90β,HSP70,and HSP27 was observed following administration of GA. We considered HMW complexes to be dimers and trimers with GA-AGEs-mediated aggregation. Cleaved caspase-3 could not be detected with WB. Furthermore,10 and 20 μg/m L GA-AGEs-BSA was 27% and 34% greater than that of control cells,respectively(P < 0.05 and P < 0.01).CONCLUSION Although intracellular GA-AGEs induce pancreatic cancer cell death,their secretion and release may promote the proliferation of other pancreatic cancer cells.
AIM To determine the possibility that diabetes mellitus promotes pancreatic ductal adenocarcinoma via glyceraldehyde(GA)-derived advanced glycation-end products(GA-AGEs).METHODS PANC-1,a human pancreatic cancer cell line,was treated with 1-4 mmol/L GA for 24 h. The cell viability and intracellular GA-AGEs were measured by WST-8 assay and slot blotting. Moreover,immunostaining of PANC-1 cells with an anti-GA-AGE antibody was performed. Western blotting(WB) was used to analyze the molecular weight of GA-AGEs. Heat shock proteins 90α,90β,70,27 and cleaved caspase-3 were analyzed by WB. In addition,PANC-1 cells were treated with GA-AGEs-bovine serum albumin(GA-AGEs-BSA),as a model of extracellular GA-AGEs,and proliferation of PANC-1 cells was measured.RESULTS In PANC-1 cells,GA induced the production of GA-AGEs and cell death in a dose-dependent manner. PANC-1 cell viability was approximately 40% with a 2 mmol/L GA treatment and decreased to almost 0% with a 4 mmol/L GA treatment(each significant difference was P < 0.01). Cells treated with 2 and 4 mmol/L GA produced 6.4 and 21.2 μg/mg protein of GA-AGEs,respectively(P <0.05 and P < 0.01). The dose-dependent production of some high-molecular-weight(HMW) complexes of HSP90β,HSP70,and HSP27 was observed following administration of GA. We considered HMW complexes to be dimers and trimers with GA-AGEs-mediated aggregation. Cleaved caspase-3 could not be detected with WB. Furthermore,10 and 20 μg/m L GA-AGEs-BSA was 27% and 34% greater than that of control cells,respectively(P < 0.05 and P < 0.01).CONCLUSION Although intracellular GA-AGEs induce pancreatic cancer cell death,their secretion and release may promote the proliferation of other pancreatic cancer cells.
AIM To determine the possibility that diabetes mellitus promotes pancreatic ductal adenocarcinoma via glyceraldehyde(GA)-derived advanced glycation-end products(GA-AGEs).METHODS PANC-1,a human pancreatic cancer cell line,was treated with 1-4 mmol/L GA for 24 h. The cell viability and intracellular GA-AGEs were measured by WST-8 assay and slot blotting. Moreover,immunostaining of PANC-1 cells with an anti-GA-AGE antibody was performed. Western blotting(WB) was used to analyze the molecular weight of GA-AGEs. Heat shock proteins 90α,90β,70,27 and cleaved caspase-3 were analyzed by WB. In addition,PANC-1 cells were treated with GA-AGEs-bovine serum albumin(GA-AGEs-BSA),as a model of extracellular GA-AGEs,and proliferation of PANC-1 cells was measured.RESULTS In PANC-1 cells,GA induced the production of GA-AGEs and cell death in a dose-dependent manner. PANC-1 cell viability was approximately 40% with a 2 mmol/L GA treatment and decreased to almost 0% with a 4 mmol/L GA treatment(each significant difference was P < 0.01). Cells treated with 2 and 4 mmol/L GA produced 6.4 and 21.2 μg/mg protein of GA-AGEs,respectively(P <0.05 and P < 0.01). The dose-dependent production of some high-molecular-weight(HMW) complexes of HSP90β,HSP70,and HSP27 was observed following administration of GA. We considered HMW complexes to be dimers and trimers with GA-AGEs-mediated aggregation. Cleaved caspase-3 could not be detected with WB. Furthermore,10 and 20 μg/m L GA-AGEs-BSA was 27% and 34% greater than that of control cells,respectively(P < 0.05 and P < 0.01).CONCLUSION Although intracellular GA-AGEs induce pancreatic cancer cell death,their secretion and release may promote the proliferation of other pancreatic cancer cells.
引文
1 Shimasaki T,Yamamoto S,Ishigaki Y,Takata T,Arisawa T,Motoo Y,Tomosug N,Minamoto T.Identification and functional analysis of an EMT-accelerating factor induced in pancreatic cancer cells by anticancer agent.Suizo 2016;31:76-84
2 Wada K,Takaori K,Traverso LW,Hruban RH,Furukawa T,Brentnall TA,Hatori T,Sano K,Takada T,Majima Y,Shimosegawa T.Clinical importance of Familial Pancreatic Cancer Registry in Japan:a report from kick-off meeting at International Symposium on Pancreas Cancer 2012.J Hepatobiliary Pancreat Sci 2013;20:557-566[PMID:23604538 DOI:10.1007/s00534-013-0611-5]
3 Sinnett-Smith J,Kisfalvi K,Kui R,Rozengurt E.Metformin inhibition of m TORC1 activation,DNA synthesis and proliferation in pancreatic cancer cells:dependence on glucose concentration and role of AMPK.Biochem Biophys Res Commun 2013;430:352-357[PMID:23159620 DOI:10.1016/j.bbrc.2012.11.010]
4 Takeuchi M.Serum Levels of Toxic AGEs(TAGE)May Be a Promising Novel Biomarker for the Onset/Progression of LifestyleRelated Diseases.Diagnostics(Basel)2016;6:pii:E23[PMID:27338481 DOI:10.3390/diagnostics6020023]
5 Bobrowski A,Spitzner M,Bethge S,Mueller-Graf F,Vollmar B,Zechner D.Risk factors for pancreatic ductal adenocarcinoma specifically stimulate pancreatic duct glands in mice.Am J Pathol2013;182:965-974[PMID:23438477]
6 Giovannucci E,Michaud D.The role of obesity and related metabolic disturbances in cancers of the colon,prostate,and pancreas.Gastroenterology 2007;132:2208-2225[PMID:17498513 DOI:10.1053/j.gastro.2007.03.050]
7 Takeuchi M,Yamagishi S.TAGE(toxic AGEs)hypothesis in various chronic diseases.Med Hypotheses 2004;63:449-452[PMID:15288366 DOI:10.1016/j.methy.2004.02.042]
8 Takino J,Nagamine K,Hori T,Sakasai-Sakai A,Takeuchi M.Contribution of the toxic advanced glycation end-productsreceptor axis in nonalcoholic steatohepatitis-related hepatocellular carcinoma.World J Hepatol 2015;7:2459-2469[PMID:26483867DOI:10.4254/wjh.v7.i23.2459]
9 Koriyama Y,Furukawa A,Muramatsu M,Takino J,Takeuchi M.Glyceraldehyde caused Alzheimer’s disease-like alterations in diagnostic marker levels in SH-SY5Y human neuroblastoma cells.Sci Rep 2015;5:13313[PMID:26304819 DOI:10.1038/srep13313]
10 Takeuchi M,Takino J,Sakasai-Sakai A,Takata T,Ueda T,Tsutsumi M,Hyogo H,Yamagishi S.Involvement of the TAGERAGE system in non-alcoholic steatohepatitis:Novel treatment strategies.World J Hepatol 2014;6:880-893[PMID:25544875DOI:10.4254/wjh.6.i12.880]
11 Takeuchi M,Sakasai-Sakai A,Takino J,Ueda T,Tsutsumi M.Toxic AGEs(TAGE)theory in the pathogenesis of NAFLD and ALD.Int J Diabetes Clin Res 2015;2:4
12 Takeuchi M,Sakasai-Sakai A,Takata T,Ueda T,Takino J,Tsutsumi M,Hyogo H,Yamagishi S.Serum levels of toxic AGEs(TAGE)may be a promising novel biomarker in development and progression of NASH.Med Hypotheses 2015;84:490-493[PMID:25697114 DOI:10.1016/j.mehy.2015.02.002]
13 Takeuchi M,Makita Z,Bucala R,Suzuki T,Koike T,Kameda Y.Immunological evidence that non-carboxymethyllysine advanced glycation end-products are produced from short chain sugars and dicarbonyl compounds in vivo.Mol Med 2000;6:114-125[PMID:10859028]
14 Jolly C,Morimoto RI.Role of the heat shock response and molecular chaperones in oncogenesis and cell death.J Natl Cancer Inst 2000;92:1564-1572[PMID:11018092]
15 Jaworek J,Leja-Szpak A.Melatonin influences pancreatic cancerogenesis.Histol Histopathol 2014;29:423-431[PMID:24258787]
16 Takino J,Kobayashi Y,Takeuchi M.The formation of intracellular glyceraldehyde-derived advanced glycation end-products and cytotoxicity.J Gastroenterol 2010;45:646-655[PMID:20084527DOI:10.1007/s00535-009-0193-9]
17 Schalkwijk CG,van Bezu J,van der Schors RC,Uchida K,Stehouwer CD,van Hinsbergh VW.Heat-shock protein 27 is a major methylglyoxal-modified protein in endothelial cells.FEBS Lett 2006;580:1565-1570[PMID:16487519]
18 Sakamoto H,Mashima T,Yamamoto K,Tsuruo T.Modulation of heat-shock protein 27(Hsp27)anti-apoptotic activity by methylglyoxal modification.J Biol Chem 2002;277:45770-45775[PMID:12226095 DOI:10.1074/jbc.M207485200]
19 Wells-Knecht KJ,Zyzak DV,Litchfield JE,Thorpe SR,Baynes JW.Mechanism of autoxidative glycosylation:identification of glyoxal and arabinose as intermediates in the autoxidative modification of proteins by glucose.Biochemistry 1995;34:3702-3709[PMID:7893666]
20 Li M,Yu X,Guo H,Sun L,Wang A,Liu Q,Wang X,Li J.Bufalin exerts antitumor effects by inducing cell cycle arrest and triggering apoptosis in pancreatic cancer cells.Tumour Biol 2014;35:2461-2471[PMID:24218335 DOI:10.1007/s13227-013-1326-6]
21 Zhu W,Li J,Wu S,Li S,Le L,Su X,Qiu P,Hu H,Yan G.Triptolide cooperates with Cisplatin to induce apoptosis in gemcitabineresistant pancreatic cancer.Pancreas 2012;41:1029-1038[PMID:22617708 DOI:10.1097/MPA.0b013e31824abdc0]
22 Jiang H,Duan B,He C,Geng S,Shen X,Zhu H,Sheng H,Yang C,Gao H.Cytoplasmic HSP90αexpression is associated with perineural invasion in pancreatic cancer.Int J Clin Exp Pathol2014;7:3305-3311[PMID:25031753]
23 Larocque K,Ovadje P,Djurdjevic S,Mehdi M,Green J,Pandey S.Novel analogue of colchicine induces selective pro-death autophagy and necrosis in human cancer cells.PLo S One 2014;9:e87064[PMID:24466327 DOI:10.1371/journal.pone.0087064]
24 Malsy M,Gebhardt K,Gruber M,Wiese C,Graf B,Bundscherer A.Effects of ketamine,s-ketamine,and MK 801 on proliferation,apoptosis,and necrosis in pancreatic cancer cells.BMC Anesthesiol2015;15:111[PMID:26219286 DOI:10.1186/s12871-015-0076-y]
25 Huggett MT,Tudzarova S,Proctor I,Loddo M,Keane MG,Stoeber K,Williams GH,Pereira SP.Cdc7 is a potent anti-cancer target in pancreatic cancer due to abrogation of the DNA origin activation checkpoint.Oncotarget 2016;7:18495-18507[PMID:26921250 DOI:10.18632/oncotarget.7611]
26 Takino J,Yamagishi S,Takeuchi M.Glycer-AGEs-RAGE signaling enhances the angiogenic potential of hepatocellular carcinoma by upregulating VEGF expression.World J Gastro-enterol 2012;18:1781-1788[PMID:22553402 DOI:10.3748/wjg.v18.i15.1781]
27 Iwamoto K,Kanno K,Hyogo H,Yamagishi S,Takeuchi M,Tazuma S,Chayama K.Advanced glycation end products enhance the proliferation and activation of hepatic stellate cells.J Gastroenterol 2008;43:298-304[PMID:18458846 DOI:10.1007/s00535-007-2152-7]
28 Sakuraoka Y,Sawada T,Okada T,Shiraki T,Miura Y,Hiraishi K,Ohsawa T,Adachi M,Takino J,Takeuchi M,Kubota K.MK615 decreases RAGE expression and inhibits TAGEinduced proliferation in hepatocellular carcinoma cells.World J Gastroenterol 2010;16:5334-5341[PMID:21072897 DOI:10 .3748/wjg.v16.i42.5334]
29 Takada M,Hirata K,Ajiki T,Suzuki Y,Kuroda Y.Expression of receptor for advanced glycation end products(RAGE)and MMP-9in human pancreatic cancer cells.Hepatogastroenterology 2004;51:928-930[PMID:15239215]
30 Takada M,Koizumi T,Toyama H,Suzuki Y,Kuroda Y.Differential expression of RAGE in human pancreatic carcinoma cells.Hepatogastroenterology 2001;48:1577-1578[PMID:11813576]
31 Takino J,Nagamine K,Takeuchi M,Hori T.In vitro identification of nonalcoholic fatty liver disease-related protein hn RNPM.World J Gastroenterol 2015;21:1784-1793[PMID:25684943 DOI:10 .3748/wjg.v21.i6.1784]
32 Takata T,Ishigaki Y,Shimasaki T,Tsuchida H,Motoo Y,Hayashi A,Tomosugi N.Characterization of proteins secreted by pancreatic cancer cells with anticancer drug treatment in vitro.Oncol Rep2012;28:1968-1976[PMID:22961650]
2 Wada K,Takaori K,Traverso LW,Hruban RH,Furukawa T,Brentnall TA,Hatori T,Sano K,Takada T,Majima Y,Shimosegawa T.Clinical importance of Familial Pancreatic Cancer Registry in Japan:a report from kick-off meeting at International Symposium on Pancreas Cancer 2012.J Hepatobiliary Pancreat Sci 2013;20:557-566[PMID:23604538 DOI:10.1007/s00534-013-0611-5]
3 Sinnett-Smith J,Kisfalvi K,Kui R,Rozengurt E.Metformin inhibition of m TORC1 activation,DNA synthesis and proliferation in pancreatic cancer cells:dependence on glucose concentration and role of AMPK.Biochem Biophys Res Commun 2013;430:352-357[PMID:23159620 DOI:10.1016/j.bbrc.2012.11.010]
4 Takeuchi M.Serum Levels of Toxic AGEs(TAGE)May Be a Promising Novel Biomarker for the Onset/Progression of LifestyleRelated Diseases.Diagnostics(Basel)2016;6:pii:E23[PMID:27338481 DOI:10.3390/diagnostics6020023]
5 Bobrowski A,Spitzner M,Bethge S,Mueller-Graf F,Vollmar B,Zechner D.Risk factors for pancreatic ductal adenocarcinoma specifically stimulate pancreatic duct glands in mice.Am J Pathol2013;182:965-974[PMID:23438477]
6 Giovannucci E,Michaud D.The role of obesity and related metabolic disturbances in cancers of the colon,prostate,and pancreas.Gastroenterology 2007;132:2208-2225[PMID:17498513 DOI:10.1053/j.gastro.2007.03.050]
7 Takeuchi M,Yamagishi S.TAGE(toxic AGEs)hypothesis in various chronic diseases.Med Hypotheses 2004;63:449-452[PMID:15288366 DOI:10.1016/j.methy.2004.02.042]
8 Takino J,Nagamine K,Hori T,Sakasai-Sakai A,Takeuchi M.Contribution of the toxic advanced glycation end-productsreceptor axis in nonalcoholic steatohepatitis-related hepatocellular carcinoma.World J Hepatol 2015;7:2459-2469[PMID:26483867DOI:10.4254/wjh.v7.i23.2459]
9 Koriyama Y,Furukawa A,Muramatsu M,Takino J,Takeuchi M.Glyceraldehyde caused Alzheimer’s disease-like alterations in diagnostic marker levels in SH-SY5Y human neuroblastoma cells.Sci Rep 2015;5:13313[PMID:26304819 DOI:10.1038/srep13313]
10 Takeuchi M,Takino J,Sakasai-Sakai A,Takata T,Ueda T,Tsutsumi M,Hyogo H,Yamagishi S.Involvement of the TAGERAGE system in non-alcoholic steatohepatitis:Novel treatment strategies.World J Hepatol 2014;6:880-893[PMID:25544875DOI:10.4254/wjh.6.i12.880]
11 Takeuchi M,Sakasai-Sakai A,Takino J,Ueda T,Tsutsumi M.Toxic AGEs(TAGE)theory in the pathogenesis of NAFLD and ALD.Int J Diabetes Clin Res 2015;2:4
12 Takeuchi M,Sakasai-Sakai A,Takata T,Ueda T,Takino J,Tsutsumi M,Hyogo H,Yamagishi S.Serum levels of toxic AGEs(TAGE)may be a promising novel biomarker in development and progression of NASH.Med Hypotheses 2015;84:490-493[PMID:25697114 DOI:10.1016/j.mehy.2015.02.002]
13 Takeuchi M,Makita Z,Bucala R,Suzuki T,Koike T,Kameda Y.Immunological evidence that non-carboxymethyllysine advanced glycation end-products are produced from short chain sugars and dicarbonyl compounds in vivo.Mol Med 2000;6:114-125[PMID:10859028]
14 Jolly C,Morimoto RI.Role of the heat shock response and molecular chaperones in oncogenesis and cell death.J Natl Cancer Inst 2000;92:1564-1572[PMID:11018092]
15 Jaworek J,Leja-Szpak A.Melatonin influences pancreatic cancerogenesis.Histol Histopathol 2014;29:423-431[PMID:24258787]
16 Takino J,Kobayashi Y,Takeuchi M.The formation of intracellular glyceraldehyde-derived advanced glycation end-products and cytotoxicity.J Gastroenterol 2010;45:646-655[PMID:20084527DOI:10.1007/s00535-009-0193-9]
17 Schalkwijk CG,van Bezu J,van der Schors RC,Uchida K,Stehouwer CD,van Hinsbergh VW.Heat-shock protein 27 is a major methylglyoxal-modified protein in endothelial cells.FEBS Lett 2006;580:1565-1570[PMID:16487519]
18 Sakamoto H,Mashima T,Yamamoto K,Tsuruo T.Modulation of heat-shock protein 27(Hsp27)anti-apoptotic activity by methylglyoxal modification.J Biol Chem 2002;277:45770-45775[PMID:12226095 DOI:10.1074/jbc.M207485200]
19 Wells-Knecht KJ,Zyzak DV,Litchfield JE,Thorpe SR,Baynes JW.Mechanism of autoxidative glycosylation:identification of glyoxal and arabinose as intermediates in the autoxidative modification of proteins by glucose.Biochemistry 1995;34:3702-3709[PMID:7893666]
20 Li M,Yu X,Guo H,Sun L,Wang A,Liu Q,Wang X,Li J.Bufalin exerts antitumor effects by inducing cell cycle arrest and triggering apoptosis in pancreatic cancer cells.Tumour Biol 2014;35:2461-2471[PMID:24218335 DOI:10.1007/s13227-013-1326-6]
21 Zhu W,Li J,Wu S,Li S,Le L,Su X,Qiu P,Hu H,Yan G.Triptolide cooperates with Cisplatin to induce apoptosis in gemcitabineresistant pancreatic cancer.Pancreas 2012;41:1029-1038[PMID:22617708 DOI:10.1097/MPA.0b013e31824abdc0]
22 Jiang H,Duan B,He C,Geng S,Shen X,Zhu H,Sheng H,Yang C,Gao H.Cytoplasmic HSP90αexpression is associated with perineural invasion in pancreatic cancer.Int J Clin Exp Pathol2014;7:3305-3311[PMID:25031753]
23 Larocque K,Ovadje P,Djurdjevic S,Mehdi M,Green J,Pandey S.Novel analogue of colchicine induces selective pro-death autophagy and necrosis in human cancer cells.PLo S One 2014;9:e87064[PMID:24466327 DOI:10.1371/journal.pone.0087064]
24 Malsy M,Gebhardt K,Gruber M,Wiese C,Graf B,Bundscherer A.Effects of ketamine,s-ketamine,and MK 801 on proliferation,apoptosis,and necrosis in pancreatic cancer cells.BMC Anesthesiol2015;15:111[PMID:26219286 DOI:10.1186/s12871-015-0076-y]
25 Huggett MT,Tudzarova S,Proctor I,Loddo M,Keane MG,Stoeber K,Williams GH,Pereira SP.Cdc7 is a potent anti-cancer target in pancreatic cancer due to abrogation of the DNA origin activation checkpoint.Oncotarget 2016;7:18495-18507[PMID:26921250 DOI:10.18632/oncotarget.7611]
26 Takino J,Yamagishi S,Takeuchi M.Glycer-AGEs-RAGE signaling enhances the angiogenic potential of hepatocellular carcinoma by upregulating VEGF expression.World J Gastro-enterol 2012;18:1781-1788[PMID:22553402 DOI:10.3748/wjg.v18.i15.1781]
27 Iwamoto K,Kanno K,Hyogo H,Yamagishi S,Takeuchi M,Tazuma S,Chayama K.Advanced glycation end products enhance the proliferation and activation of hepatic stellate cells.J Gastroenterol 2008;43:298-304[PMID:18458846 DOI:10.1007/s00535-007-2152-7]
28 Sakuraoka Y,Sawada T,Okada T,Shiraki T,Miura Y,Hiraishi K,Ohsawa T,Adachi M,Takino J,Takeuchi M,Kubota K.MK615 decreases RAGE expression and inhibits TAGEinduced proliferation in hepatocellular carcinoma cells.World J Gastroenterol 2010;16:5334-5341[PMID:21072897 DOI:10 .3748/wjg.v16.i42.5334]
29 Takada M,Hirata K,Ajiki T,Suzuki Y,Kuroda Y.Expression of receptor for advanced glycation end products(RAGE)and MMP-9in human pancreatic cancer cells.Hepatogastroenterology 2004;51:928-930[PMID:15239215]
30 Takada M,Koizumi T,Toyama H,Suzuki Y,Kuroda Y.Differential expression of RAGE in human pancreatic carcinoma cells.Hepatogastroenterology 2001;48:1577-1578[PMID:11813576]
31 Takino J,Nagamine K,Takeuchi M,Hori T.In vitro identification of nonalcoholic fatty liver disease-related protein hn RNPM.World J Gastroenterol 2015;21:1784-1793[PMID:25684943 DOI:10 .3748/wjg.v21.i6.1784]
32 Takata T,Ishigaki Y,Shimasaki T,Tsuchida H,Motoo Y,Hayashi A,Tomosugi N.Characterization of proteins secreted by pancreatic cancer cells with anticancer drug treatment in vitro.Oncol Rep2012;28:1968-1976[PMID:22961650]