A study has found that some miRNAs in diet are not taken up into mice tissues. But how comprehensive are the results?

In 2012, Chinese researchers published a paper showing that minute fragments of plant genetic material – so-called microRNA or miRNA molecules – from rice were taken up into the blood and organs of mice that ate the rice, where they regulated the expression of certain genes and affected the animals’ physiological processes.

The paper caused a panic in GMO industry circles, prompting a number of attempted “damage control” papers. These included one paper by employees of Monsanto and a biopharmaceuticals company and another by Monsanto employees.

The first paper did not find uptake of rice miRNAs in the blood plasma and liver of mice or any physiological changes that could indicate gene regulation from the ingested miRNAs.

The second paper concluded that miRNAs found in animals can be an artefact of the sequencing methodology used to detect them and that accumulation of plant miRNA from the diet is “not a common faculty among animals”. Not that lack of common-ness is reassuring: plane crashes are not common but their effects are nonetheless serious.

It should be noted that neither paper disproved the findings of the Chinese researchers, who suggested in a reply to the first Monsanto paper that the Monsanto authors’ failure to find the miRNAs may have been due to weaknesses in the sequencing method they used.

Now a new paper published in the Journal of Biological Chemistry once again gives a reassuring message about the fate of dietary miRNAs. The authors looked at two types of miRNA present in the maternal milk of mice and investigated what happened to them after the milk was ingested by the mouse pups. One type of miRNA was used as a positive control to test the sensitivity of the experiment. The authors found that these two milk miRNAs were not taken up into the mouse pups’ tissues or blood, but were degraded by the digestive system.

So far so good. But then, rather strangely, the authors extrapolated from a finding on just two types of miRNA out of 500 that they located in milk, to conclude that nutritionally derived miRNAs in general “are unlikely to cross the intestinal barrier and influence gene expression”.

A news release from the Swiss Federal Institute of Technology in Zurich (ETH Zurich), where the work was carried out, followed the authors in this incorrect generalisation in a press release titled, “MicroRNAs are digested, not absorbed”. The press release said of the research, “Experiments using mouse models show that the dietary uptake of microRNAs is barely significant, and certainly insufficient to affect physiological functions. Moreover, the microRNA molecules are broken down in the small intestine.”

Should we be reassured? We asked Dr Sarah Agapito, a research scientist at Genøk, Norway, who has co-authored papers on miRNAs, to comment on the new study. She said:

“The authors found an average of 500 miRNAs present in milk extracted from the mouse pups' stomachs. They decided to investigate just two of those miRNAs and apparently these two miRNAs could not be found in intestinal epithelium, blood, liver and spleen of pups fed on this milk. But we know that there is selective packaging of miRNAs into microvesicles, which are thought to protect certain miRNAs and facilitate their transport to target tissues, where they may be preferentially taken up by those tissues. Thus each of those 500 miRNAs could be differently processed in the cells of an organism fed on them. This investigation of two miRNAs cannot rule out the possibility that other miRNAs could be taken up into the blood or tissues of the test animals.”

Another paper on miRNA with industry involvement

Lead author of the new paper, Markus Stoffel, lists in the conflicts of interest section that he is a member of the scientific advisory board of Regulus Therapeutics and Alnylam Pharmaceuticals, pharmaceutical companies that develop miRNA drugs.

This doesn’t of course prove that Stoffel and colleagues’ findings are wrong or even questionable. They add valuable knowledge to the current murkiness surrounding the fate of ingested miRNAs.

It does, however, show that he has an interest in downplaying the alarm that has spread through some scientific circles about the possible off-target effects of miRNA-based GM products (specifically: they might silence our genes). Might this explain why he incorrectly generalised his findings from a very limited study on just two types of miRNA to make a reassuring conclusion about miRNAs in general?
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Uptake and function studies of maternal milk-derived microRNAs

Alexandra C. Title, Rémy Denzler and Markus Stoffel
J. Biol. Chem. 2015, Published Aug 3, 2015
http://www.jbc.org/content/early/2015/08/03/jbc.M115.676734.abstract
(open access)

Background: It is unclear whether maternal milk microRNAs are taken up by offspring.
Results: Milk microRNAs are not taken up into murine offspring tissues or blood, but are degraded by the digestive system.
Conclusion: It is unlikely that milk microRNAs function through canonical microRNA silencing.
Significance: Nutritionally derived microRNAs are unlikely to cross the intestinal barrier and influence gene expression.

ABSTRACT
MicroRNAs (miRNAs) are important regulators of cell-autonomous gene expression that influence many biological processes. They are also released from cells and are present in virtually all body fluids, including blood, urine, saliva, sweat, and milk. The functional role of nutritionally obtained extracellular miRNAs is controversial, and irrefutable demonstration of exogenous miRNA uptake by cells and canonical miRNA function is still lacking. Here we show that miRNAs are present at high levels in milk of lactating mice. To investigate intestinal uptake of miRNAs in newborn mice, we employed genetic models in which newborn miR-375 and miR-200c/141 knockout mice received milk from wildtype foster mothers. Analysis of intestinal epithelium, blood, liver and spleen revealed no evidence for miRNA uptake. miR-375 levels in hepatocytes were at the limit of detection and remained orders of magnitude below the threshold for target gene regulation (between 1000 and 10,000 copies/cell). Furthermore, our study revealed rapid degradation of milk miRNAs in the intestinal fluid. Together, our results indicate a nutritional rather than gene regulatory role of miRNAs in milk of newborn mice.