High glucose enhances progression of cholangiocarcinoma cells via STAT3 activation

Abstract

Epidemiological studies have indicated diabetes mellitus (DM) as a risk of cholangiocarcinoma (CCA), however, the effects and mechanisms of high glucose on progression of CCA remain unclear. This study reports for the first time of the enhancing effects of high glucose on aggressive phenotypes of CCA cells via STAT3 activation. CCA cells cultured in high glucose media exerted significantly higher rates of cell proliferation, adhesion, migration and invasion than those cultured in normal glucose. The phosphokinase array revealed STAT3 as the dominant signal activated in response to high glucose. Increased nuclear STAT3, p-STAT3 and its downstream target proteins, cyclin D1, vimentin and MMP2, were shown to be underling mechanisms of high glucose stimulation. The link of high glucose and STAT3 activation was confirmed in tumor tissues from CCA patients with DM that exhibited higher STAT3 activation than those without DM. Moreover, the levels of STAT3 activation were correlated with the levels of blood glucose. Finally, decreasing the level of glucose or using a STAT3 inhibitor could reduce the effects of high glucose. These findings suggest that controlling blood glucose or using a STAT3 inhibitor as an alternative approach may improve the therapeutic outcome of CCA patients with DM.

Metformin Exerts Antiproliferative and Anti-metastatic Effects Against Cholangiocarcinoma Cells by Targeting STAT3 and NF-ĸB

“…Cells were then treated with 5, 10, 20, 40 mM metformin for 48 h or for different times. The number of viable cells was determined using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay as described in our previous study (14).…”

“…CCA cells were pretreated with 10 mM metformin for 24 h prior to migration and invasion assays according to our previous report (14). Cells were allowed to migrate in the absence of metformin for 12 h for KKU-M213 and 24 h for KKU-M214 cells.…”

“…CCA cells were treated with 10, 20, 40 mM metformin for 48 h whereas those for protein markers of metastasis were treated with 10 mM metformin for 24 h. Cells were lysed with NP-40 lysis buffer containing protease and phosphatase inhibitor cocktail (Roche, Mannheim, Germany). Cell lysates were prepared, subjected to sodium dodecyl sulphate -polyacrylamide gel electrophoresis and transferred to a Hybond™-polyvinylidene fluoride (PVDF) membrane as mentioned previously (14). The membrane was probed with each primary antibody at 4˚C overnight and with horseradish peroxidaseconjugated secondary antibody for 1 h at room temperature.…”

“…CCA cells (8×10 3 cells/well) were cultured in Matrigel pre-coated slide chambers overnight at 37˚C with 5% CO 2 . The cells were treated 10 and 20 mM metformin for 24 h. Immunofluorescent staining was performed and recorded as described previously (14). Fixed cells were incubated with 1:100 anti-STAT3 or anti-p65 antibodies at 4˚C overnight.…”

Low Sugar Diet: A New Tool in War Against Tumorigenesis

“…Activation of STAT3, increase in level of p-STAT3 and nuclear translocation of p-STAT3 along with up-regulations of cyclin D1, vimentin, and matrix metalloproteinase 2 were responsible for growth progression in response to high glucose condition. These observations were verified not only in CCA cell lines but were also found to be true in CCA patient tissues [7]. High glucose (HG) also encouraged migration and Invasion of CCA cells and the effect was found to be more pronounced in the highly metastatic sublines.…”