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Role of microRNA in zinc depletion-induced growth inhibition in mouse fibroblast 3T3 cells

University of British Columbia

Zinc, an essential trace element for humans, exerts many physiological functions, including its indispensible role in growth. Dietary zinc deficiency in children results in growth retardation. However, the mechanism whereby zinc regulates growth remains unclear. MicroRNAs are a group of newly discovered, small, non-coding RNAs and have been demonstrated to play a regulatory role in cell proliferation. The expression of microRNA and process of producing mature microRNA can be influenced by cellular and tissue zinc status. Thus, the hypothesis for my research is that microRNA plays a role in mediating zinc-dependent cell proliferation. The overall objective was to determine whether microRNAs are involved in zinc-dependent cell proliferation in mouse fibroblast 3T3 cells. 3T3 cells were cultured in Dulbecco’s Modified Eagle Medium with 10% fetal bovine serum (FBS) for 3 days. To deplete intracellular zinc, cells were cultured in the same media containing 5% (v/v) dimethyl sulfoxide (DMSO, control) or N,N,N’,N’- tetrakis(2-pyridylmethyl)ethylenediamine (TPEN, 2.5 μM) for 24 or 48 h with or without induction of quiescence (aphidicolin, 0.5 μg/ml; 24 h). To establish zinc-dependent effect, zinc was replenished at the final concentration of 0 (TPEN only), 1.25, 2.5 or 5 μM for 24 or 48 h. Cell proliferation was measured by cell cycle analysis using flow cytometry. MicroRNA expression profile was assessed by microRNA microarray. In the absence of quiescence induction, zinc-depletion for 24 or 48 h inhibited cell proliferation by 10.4% and 16.0% compared to control. In the presence of quiescence induction, zinc-depletion inhibited cell proliferation by 52.9%. Regardless of the status of quiescence, zinc replenishment at 1.25 μM nearly brought cell proliferation back to the level observed in the DMSO control, showing a zinc-dependent cell proliferation in 3T3 cells. Zinc-depletion increased the abundance of miR-132-3p, miR-212-3, and let-7e-3p, while zinc replenishment brought back the abundance of these three microRNAs to the level observed in the DMSO control. Interestingly, zinc-depletion decreased the abundance of miR-145b, and its abundance was increased after zinc replenishment. Overall, it appeared that microRNA played a role in zinc-mediated growth regulation in 3T3 cells; however, this role of microRNA remains to be affirmed by further investigation.

Text

2014-08-28

Attribution-NonCommercial-NoDerivs 2.5 Canada

http://hdl.handle.net/2429/50254

Human Nutrition

University of British Columbia

UBCV

http://creativecommons.org/licenses/by-nc-nd/2.5/ca/