Abstract
Background
Colorectal cancer is a worldwide problem which has been associated with changes in diet and lifestyle pattern. As a result of colonic fermentation of dietary fibres, short chain free fatty acids are generated which activate Free Fatty Acid Receptors 2 and 3 (FFAR2 and FFAR3). FFAR2 and FFAR3 genes are abundantly expressed in colonic epithelium and play an important role in the metabolic homeostasis of colonic epithelial cells. Earlier studies point to the involvement of FFAR2 in colorectal carcinogenesis.
Methods
Transcriptome analysis console was used to analyse microarray data from patients and cell lines. We employed shRNA mediated down regulation of FFAR2 and FFAR3 genes which was assessed using qRT-PCR. Assays for glucose uptake and cAMP generation was done along with immunofluorescence studies. For measuring cell proliferation, we employed real time electrical impedance based assay available from xCelligence.
Results
Microarray data analysis of colorectal cancer patient samples showed a significant down regulation of FFAR2 gene expression. This prompted us to study the FFAR2 in colorectal cancer. Since, FFAR3 shares significant structural and functional homology with FFAR2, we knocked down both these receptors in colorectal cancer cell line HCT 116. These modified cell lines exhibited higher proliferation rate and were found to have increased glucose uptake as well as increased level of GLUT1. Since, FFAR2 and FFAR3 signal through G protein subunit (Gαi), knockdown of these receptors was associated with increased cAMP. Inhibition of PKA did not alter the growth and proliferation of these cells indicating a mechanism independent of cAMP/PKA pathway. Conclusion: Our results suggest role of FFAR2/FFAR3 genes in increased proliferation of colon cancer cells via enhanced glucose uptake and exclude the role of protein kinase A mediated cAMP signalling. Alternate pathways could be involved that would ultimately result in increased cell proliferation as a result of down regulated FFAR2/FFAR3 genes. This study paves the way to understand the mechanism of action of short chain free fatty acid receptors in colorectal cancer.
Release of endothelial cell lipoprotein lipase by plasma lipoproteins and free fatty acids
