Author: Karen Joy Humphreys
Humphreys, Karen Joy, 2012 Dietary regulation of microRNA expression in colorectal cells, Flinders University, School of Medicine
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Colorectal cancer (CRC) development is associated with epigenetic modifications, including DNA methylation changes, altered histone modification patterns, and dysregulated microRNA (miRNA) expression. While some dietary compounds can alter colorectal cell behaviour through epigenetic mechanisms, their role in modifying miRNA expression in CRC cells and normal colorectal tissue has been less studied. The diet-derived compound butyrate, with its known role in histone modification, is a plausible candidate for altering miRNA expression. This study examined dietary regulation of miRNA expression in colorectal cells, and explored the role of butyrate and other histone deacetylase inhibitors (HDIs) in modulating CRC risk through altered miRNA expression. The down-stream consequences of these miRNA changes, and the roles of miRNAs in the context of the anti-proliferative effects of HDIs, were determined. In addition to exploring the action of butyrate, a potentially protective dietary component, the study also investigated whether factors that possibly increase CRC risk, such as high red meat intake, alter miRNA expression. In vitro, butyrate and other HDIs altered levels of some miRNAs that are dysregulated in CRC, including the oncogenic miR-17-92 miRNA cluster which is over-expressed in CRC. Butyrate decreased miR-17-92 miRNA levels in CRC cells, with a corresponding increase in expression of miR-17-92 targets, including cell cycle inhibitors and pro-apoptotic genes. Mechanisms for this decrease included changes in regulators of miR-17-92 host gene transcription, and altered histone acetylation and methylation patterns centred around the transcription start site and promoter of the miR-17-92 host gene. Decreased miR-17-92 expression may be partly responsible for the anti-proliferative effects of HDIs, with introduction of miR-17-92 cluster miRNA mimics reversing this effect and decreasing target gene transcript levels. Of the cluster members, miR-19a and miR-19b were primarily responsible for promoting proliferation, while in a novel finding, miR-18a acted in opposition to other members to decrease growth. Two pro-proliferative genes, NEDD9 and CDK19, were identified as novel miR-18a targets. This study presents the first evidence of competing roles for miR-17-92 cluster members, in the context of HDI-induced changes in CRC. miR-18a may play a homeostatic role in containing the oncogenic effects of the entire cluster, but may be selectively decreased in CRC compared with other cluster members. In addition to the capacity of butyrate to reverse the dysregulation of miR-17-92 miRNAs in CRC cells in vitro, this action was demonstrated with resistant starch supplementation in vivo, in rectal biopsies from healthy human volunteers exposed to high red meat levels. High red meat intake raised levels of miRNAs with oncogenic potential, particularly miR-17-92 cluster miRNAs and miR-21. Resistant starch supplementation raised faecal butyrate concentrations, and decreased miR-17-92 cluster miRNAs to baseline levels. In vivo modulation of miRNAs in colorectal cells by dietary compounds has not previously been demonstrated in humans. Regulation of miRNA expression demonstrates a plausible mechanism to explain some of the chemo-protective effects of butyrate, and potentially carcinogenic properties of other dietary components. Understanding how dietary compounds alter miRNA expression, and how miRNAs modulate the action of HDIs, may provide new opportunities for CRC therapies and prevention strategies.
Keywords: histone deacetylase inhibitors,butyrate,microRNAs,miR-17-92,colorectal cancer,epigenetic
Subject: Nutrition and Dietetics thesis, Medicine thesis
Thesis type: Doctor of Philosophy
School: School of Medicine
Supervisor: Dr Michael Michael