Background & Aims: Diabetes mellitus (DM) has been identified as a risk factor for a variety of malignancies, including pancreatic cancer (PaC). Many factors may promote PaC in DM, including hyperglycemia itself, dyslipidemia, chronic inflammation and insulin resistance/hyperinsulinemia, but the mechanistic link is unknown. Previous studies have demonstrated a tumor-promoting role for RAGE, the receptor for advanced glycation endproducts (AGEs), which levels are increased in diabetes. Here we test the hypothesis of a promoting role of AGEs in PaC through RAGE activation. Methods: We investigated the effects of the AGE Nε-carboxymethyllysine (CML) in human pancreatic ductal adenocarcinoma (PDA) cell lines and in a mouse model of Kras-driven PaC interbred with a bioluminescent model of proliferation. PaC–prone KCM (LSL-KrasG12D/+;Pdx-1-Cre;MITO) mice, which spontaneously develop autochthonous lethal PaC, were generated by interbreeding mutated KCM transgenic mice with MITO-Luc reporter mice. In this model, the promoter of the luciferase reporter is under the control of the nuclear factor (NF)-Y, which is specifically expressed in proliferating cells. Therefore, this animal model gave us the chance to monitor the tumor growth in vivo by bioluminescence imaging. Results were confirmed by histology examination. In vitro results: In the range of 10-100 μg/mL, CML promoted cell growth and RAGE expression (~1,15 to 1,4 times, p<0.01) in a concentration- and time-dependent manner in two PDA cell lines. At variance, normal human pancreatic duct epithelial cells showed a positive response to CML only at the lowest concentration (1 μg/mL) tested. Interestingly, the different proliferative response to CML observed in different cell lines was positively associated with basal expression levels of RAGE: i.e., the higher was the basal expression of RAGE of the cell line, the larger was the range of CML concentrations capable of providing an additional proliferative stimulus at 24 h. Mechanistically, the activation of the AGE/RAGE axis in PDA cells increased NK-κB p65 protein level and activation (~1,5 and 3 times, respectively, p<0,01), STAT3 phosphorilation (~3 times, p<0,01), and the transcriptional activity of its targets genes PIM1 and NFATc1 (~3 and 1,5 times, respectively, p<0.01), which have been involved in PaC promotion. These effects were completely counteracted by RAP (RAGE antagonist peptide), except for PIM1 expression and STAT3 phosphorilation, which were only partially, but significantly (more than halved, p<0,01), reduced. In vivo results: Five-week old KCM mice were daily treated intraperitoneally (i.p.) with 30 μg of CML-modified (CML mice) or native mouse serum albumin (MSA, vehicle; Ctr mice). At 11 weeks of age (6 weeks of treatment), more than half of CML mice (six of eleven) showed positive abdominal imaging and about 1/4 (three of eleven) had a palpable abdominal mass. At the histological analysis, invasive PaC was confirmed in eight of eleven (72.7%) CML mice, compared with only one of eleven (9.1%, Fisher's exact test p<0,001) Ctr mice. IHC analysis for the distribution of RAGE in pancreatic sections of CML mice showed intense staining in neoplastic precursor lesions PanINs, but not of the surrounding acinar tissue. Immunoreactivity for RAGE was also observed in cells of duct-like structures in well-differentiated regions of the tumor, but not in surrounding undifferentiated areas. Fainter RAGE staining in PaiINs lesions was observed in Ctr mice. Conclusions: The AGE/RAGE axis modulates development and progression of PaC. The finding that AGEs accelerate PaC development from PanINs lesions might explain the increased risk conferred by DM and identifies AGEs as possible therapeutic target for risk reduction in diabetic patients.
Background & Aims: Diabetes mellitus (DM) has been identified as a risk factor for a variety of malignancies, including pancreatic cancer (PaC). Many factors may promote PaC in DM, including hyperglycemia itself, dyslipidemia, chronic inflammation and insulin resistance/hyperinsulinemia, but the mechanistic link is unknown. Previous studies have demonstrated a tumor-promoting role for RAGE, the receptor for advanced glycation endproducts (AGEs), which levels are increased in diabetes. Here we test the hypothesis of a promoting role of AGEs in PaC through RAGE activation. Methods: We investigated the effects of the AGE Nε-carboxymethyllysine (CML) in human pancreatic ductal adenocarcinoma (PDA) cell lines and in a mouse model of Kras-driven PaC interbred with a bioluminescent model of proliferation. PaC–prone KCM (LSL-KrasG12D/+;Pdx-1-Cre;MITO) mice, which spontaneously develop autochthonous lethal PaC, were generated by interbreeding mutated KCM transgenic mice with MITO-Luc reporter mice. In this model, the promoter of the luciferase reporter is under the control of the nuclear factor (NF)-Y, which is specifically expressed in proliferating cells. Therefore, this animal model gave us the chance to monitor the tumor growth in vivo by bioluminescence imaging. Results were confirmed by histology examination. In vitro results: In the range of 10-100 μg/mL, CML promoted cell growth and RAGE expression (~1,15 to 1,4 times, p<0.01) in a concentration- and time-dependent manner in two PDA cell lines. At variance, normal human pancreatic duct epithelial cells showed a positive response to CML only at the lowest concentration (1 μg/mL) tested. Interestingly, the different proliferative response to CML observed in different cell lines was positively associated with basal expression levels of RAGE: i.e., the higher was the basal expression of RAGE of the cell line, the larger was the range of CML concentrations capable of providing an additional proliferative stimulus at 24 h. Mechanistically, the activation of the AGE/RAGE axis in PDA cells increased NK-κB p65 protein level and activation (~1,5 and 3 times, respectively, p<0,01), STAT3 phosphorilation (~3 times, p<0,01), and the transcriptional activity of its targets genes PIM1 and NFATc1 (~3 and 1,5 times, respectively, p<0.01), which have been involved in PaC promotion. These effects were completely counteracted by RAP (RAGE antagonist peptide), except for PIM1 expression and STAT3 phosphorilation, which were only partially, but significantly (more than halved, p<0,01), reduced. In vivo results: Five-week old KCM mice were daily treated intraperitoneally (i.p.) with 30 μg of CML-modified (CML mice) or native mouse serum albumin (MSA, vehicle; Ctr mice). At 11 weeks of age (6 weeks of treatment), more than half of CML mice (six of eleven) showed positive abdominal imaging and about 1/4 (three of eleven) had a palpable abdominal mass. At the histological analysis, invasive PaC was confirmed in eight of eleven (72.7%) CML mice, compared with only one of eleven (9.1%, Fisher's exact test p<0,001) Ctr mice. IHC analysis for the distribution of RAGE in pancreatic sections of CML mice showed intense staining in neoplastic precursor lesions PanINs, but not of the surrounding acinar tissue. Immunoreactivity for RAGE was also observed in cells of duct-like structures in well-differentiated regions of the tumor, but not in surrounding undifferentiated areas. Fainter RAGE staining in PaiINs lesions was observed in Ctr mice. Conclusions: The AGE/RAGE axis modulates development and progression of PaC. The finding that AGEs accelerate PaC development from PanINs lesions might explain the increased risk conferred by DM and identifies AGEs as possible therapeutic target for risk reduction in diabetic patients.
Diabetes and pancreatic cancer: role of advanced glycation endproducts (AGEs) in tumor progression via RAGE activation / Menini, Stefano; Iacobini, Carla; BLASETTI FANTAUZZI, Claudia; Delatouliere, Luisa; Manni, Isabella; Cappello, Paola; Novelli, Francesco; Pugliese, Giuseppe. - CD-ROM. - ??:(2017), pp. ??-??. (Intervento presentato al convegno 39 Congresso Nazionale Società Italiana di Endocrinologia (SIE) tenutosi a Palazzo dei Congressi, Roma nel 21-24 giugno).
Diabetes and pancreatic cancer: role of advanced glycation endproducts (AGEs) in tumor progression via RAGE activation
MENINI, Stefano;IACOBINI, carla;BLASETTI FANTAUZZI, CLAUDIA;PUGLIESE, Giuseppe
2017
Abstract
Background & Aims: Diabetes mellitus (DM) has been identified as a risk factor for a variety of malignancies, including pancreatic cancer (PaC). Many factors may promote PaC in DM, including hyperglycemia itself, dyslipidemia, chronic inflammation and insulin resistance/hyperinsulinemia, but the mechanistic link is unknown. Previous studies have demonstrated a tumor-promoting role for RAGE, the receptor for advanced glycation endproducts (AGEs), which levels are increased in diabetes. Here we test the hypothesis of a promoting role of AGEs in PaC through RAGE activation. Methods: We investigated the effects of the AGE Nε-carboxymethyllysine (CML) in human pancreatic ductal adenocarcinoma (PDA) cell lines and in a mouse model of Kras-driven PaC interbred with a bioluminescent model of proliferation. PaC–prone KCM (LSL-KrasG12D/+;Pdx-1-Cre;MITO) mice, which spontaneously develop autochthonous lethal PaC, were generated by interbreeding mutated KCM transgenic mice with MITO-Luc reporter mice. In this model, the promoter of the luciferase reporter is under the control of the nuclear factor (NF)-Y, which is specifically expressed in proliferating cells. Therefore, this animal model gave us the chance to monitor the tumor growth in vivo by bioluminescence imaging. Results were confirmed by histology examination. In vitro results: In the range of 10-100 μg/mL, CML promoted cell growth and RAGE expression (~1,15 to 1,4 times, p<0.01) in a concentration- and time-dependent manner in two PDA cell lines. At variance, normal human pancreatic duct epithelial cells showed a positive response to CML only at the lowest concentration (1 μg/mL) tested. Interestingly, the different proliferative response to CML observed in different cell lines was positively associated with basal expression levels of RAGE: i.e., the higher was the basal expression of RAGE of the cell line, the larger was the range of CML concentrations capable of providing an additional proliferative stimulus at 24 h. Mechanistically, the activation of the AGE/RAGE axis in PDA cells increased NK-κB p65 protein level and activation (~1,5 and 3 times, respectively, p<0,01), STAT3 phosphorilation (~3 times, p<0,01), and the transcriptional activity of its targets genes PIM1 and NFATc1 (~3 and 1,5 times, respectively, p<0.01), which have been involved in PaC promotion. These effects were completely counteracted by RAP (RAGE antagonist peptide), except for PIM1 expression and STAT3 phosphorilation, which were only partially, but significantly (more than halved, p<0,01), reduced. In vivo results: Five-week old KCM mice were daily treated intraperitoneally (i.p.) with 30 μg of CML-modified (CML mice) or native mouse serum albumin (MSA, vehicle; Ctr mice). At 11 weeks of age (6 weeks of treatment), more than half of CML mice (six of eleven) showed positive abdominal imaging and about 1/4 (three of eleven) had a palpable abdominal mass. At the histological analysis, invasive PaC was confirmed in eight of eleven (72.7%) CML mice, compared with only one of eleven (9.1%, Fisher's exact test p<0,001) Ctr mice. IHC analysis for the distribution of RAGE in pancreatic sections of CML mice showed intense staining in neoplastic precursor lesions PanINs, but not of the surrounding acinar tissue. Immunoreactivity for RAGE was also observed in cells of duct-like structures in well-differentiated regions of the tumor, but not in surrounding undifferentiated areas. Fainter RAGE staining in PaiINs lesions was observed in Ctr mice. Conclusions: The AGE/RAGE axis modulates development and progression of PaC. The finding that AGEs accelerate PaC development from PanINs lesions might explain the increased risk conferred by DM and identifies AGEs as possible therapeutic target for risk reduction in diabetic patients.I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
