The carcinogenic potential of trans-fatty acids: investigation of effects on the miRNA expression profile of human colon cancer cells

The carcinogenic potential of trans-fatty acids: Investigation of effects on the miRNA expression profile of human colon cancer cells Trans fatty acids (TFA) are unsaturated fatty acids with at least one double-bond in trans configuration. They are generated during the industrial hardening of plant oils and refinement processes at high temperatures or formed by bacterial metabolism in the rumen of ruminant animals. A clear correlation between the consumption of TFA and increased risk of coronary heart disease was shown while a potential impact on the genesis of colorectal cancer (CRC) is still controversially discussed. MiRNAs are small, non-coding RNAs that regulate gene expression on a post-transcriptional level. By regulating target mRNAs they can either act as oncogenes or tumor suppressors. MiRNAs are differentially expressed in a multitude of tumors and can act as valuable tools in the diagnosis, staging and potentially therapy of different types of cancer. The aim of this study was to analyze potential cis/trans specific fatty acid-induced changes of the miRNA expression profile in human colon cancer cells to investigate their possible role in colon carcinogenesis. In this study, 16 different C18 isomers with double bonds in trans- and/or cis-configuration were investigated. Caco-2 colon cancer cells were incubated for 24 h in the presence of different fatty acids (FA). Appropriate noncytotoxic concentrations were determined prior to the main experiment using CTB assays. Subsequently, the expression of 84 cancer-associated miRNAs was analyzed by qRT-PCR using the Human Cancer PathwayFinder miScript qRT-PCR Array (Qiagen, Hilden). 66 of the 84 investigated miRNAs were significantly deregulated after FA exposure. In particular, hsa-miR-32-5p and hsa-miR-146a-5p were most strongly regulated. Hsa-miR-32-5p was downregulated by eight FA and hsa-miR-146a-5p was upregulated by eight of the investigated FA but not comparably strong. After review of the literature for possible targets of the significantly regulated miRNAs, different proteins of the BCL-2 pathway were chosen for further investigation. The expression of BIM (BCL2L11), BCL-2 and BCL-XL was investigated on the mRNA level via SYBR Green qRT-PCR. The study provides first evidence that the fatty acid-induced downregulation of miR-32 correlates with an upregulation of BIM. Thus, the data support the hypothesis that FA have an impact on the induction of apoptosis in Caco-2 cells via downregulation of miR-32. The fatty acid-induced effects, however, were not dependent on the presence of trans double bonds and cis isomers induced comparable effects.