Antitumor effect of metformin in esophageal cancer: In vitro study

Abstract

Recent studies suggest that metformin, which is a member of the biguanide family and commonly used as an oral anti-hyperglycemic agent, may reduce cancer risk and improve prognosis of numerous types of cancer. However, the mechanisms underlying the antitumor effect of metformin on esophageal cancer remain unknown. The goal of the present study was to evaluate the effects of metformin on the proliferation of human ESCC in vitro, and to study changes in the expression profile of microRNAs (miRNAs), since miRNAs have previously been associated with the antitumor effects of metformin in other human cancers. The human ESCC cell lines T.T, KYSE30 and KYSE70 were used to study the effects of metformin on human ESCC in vitro. In addition, we used miRNA array tips to explore the differences between miRNAs in KYSE30 cells with and without metformin treatment. Metformin inhibited the proliferation of T.T, KYSE30 and KYSE70 cells in vitro. Metformin blocked the cell cycle in G0/G1 in vitro. This blockade was accompanied by a strong decrease of G1 cyclins, especially cyclin D1, as well as decreases in cyclin-dependent kinase (Cdk)4, Cdk6 and phosphorylated retinoblastoma protein (Rb). In addition, the expression of miRNAs was markedly altered with the treatment of metformin in vitro. Metformin inhibited the growth of three ESCC cell lines, and this inhibition may have involved reductions in cyclin D1, Cdk4 and Cdk6.

Mechanism of gemcitabine-induced suppression of human cholangiocellular carcinoma cell growth

“…Cell proliferation assays were conducted using CCK-8 according to the manufacturer's instructions, as in our previous studies (9,11,12). Each cell line (1x10 4 ) was seeded into a well of a 96-well plate and cultured in 100 µl of each culture medium.…”

“…Lysates were collected according to the methods described in our previous studies (9,11,12). All steps were performed at 4˚C.…”

“…Aberrant miRNA expression is a common feature of various human malignancies (9), and some studies have demonstrated that specific miRNAs are expressed in CCC cells compared with non-malignant cells (10). In addition, many studies, including our own, have reported that miRNAs play an important role in the antitumor effect of anticancer therapeutics (9,11,12). Therefore, the present study was undertaken to identify the miRNAs associated with the antitumor effects of gemcitabine in CCC.…”

“…HuCCT-1 cells (1.0x10 6 cells in a 6-well plate dish) were treated with 0.1 µg/ml gemcitabine or without gemcitabine for 24-72 h. After treatment, the cells were harvested and fixed in 80% ethanol, washed with PBS, and stored at -20˚C until flow cytometric analysis was conducted as in our previous studies (9,11,12). Prior to analysis, the cells were washed in cold PbS and resuspended in 100 µl of PbS and 10 µl of RNase A solution (250 µg/ml), followed by incubation for 30 min at 37˚C.…”

“…AAH-ANG-1) was purchased from Raybiotech Inc. (Norcross, GA, USA). The assay for the array was performed according to the manu facturer's instructions as in our previous studies (9,11,12). Briefly, the angiogenesis antibody membranes were incubated in blocking buffer for 30 min.…”