Chapter 169
MicroRNAs in Food
Randall Munroe, the nerd cartoonist of xkcd.com, reminds us in his cartoon “Every gathering is really a family reunion” that, in the end, we are all related to one another. If you go back far enough, we all have a common ancestor, all the way back to the very first Homo sapiens from whom we all descend. The cartoon shows a party scene with stick figures labeled “me,” “second cousin,” “14th cousin,” and “35th cousin,” plus a cat called “17,000,000th cousin.” Yes, if you go back far enough, you and the kitty really do have a true common ancestor of flesh and blood. In the cartoon there is also a houseplant labeled “50,000,000,000th cousin.” Using a “molecular clock,” based on corresponding DNA deviations, it has been estimated that plants and animals diverged about 1.576 billion years ago plus/minus 88 million. So you too, kitty, have a common ancestor with the ficus. Indeed: a family reunion.
The realization that microRNAs are ubiquitous and active in plants followed soon after they were discovered in animals. Cotton plants, for example, use microRNAs to switch off virulence genes of a pathogenic fungus. How might plant microRNAs interact with other cross-border actions—on us? Just as we share numerous microRNAs with other animals, certain microRNA sequences in plants overlap so strongly with those of animals that scientists suspect they are one and the same microRNA that has been conserved over 1.5 billion years of evolution. Be that as it may, if you compare the sequences of plant microRNAs with human messenger RNA, there appear to be at least 1000 different human genes that plant microRNAs could have in their sights.
Plant-based diets contain thousands of biologically active microRNAs. While the scientific community in the past attributed the benefits of fruits, vegetables, and medicinal plants to their phytonutrients, it may well be that microRNAs also play a role. Isolated phytonutrients often could not fully reproduce the effects of the whole foods from which they were extracted. That was attributed to the synergistic symphony of the various components. As we have seen, phytonutrients such as polyphenols influence our physiology, among other ways, by manipulating microRNA expression, but perhaps the genes are being switched off directly by plant microRNAs.
Research into the potential of cross-kingdom gene regulation with “xeno-microRNAs” from plants is currently considered one of the most exciting research areas in all of science. In the broadest sense, the concept of cross-kingdom genetic manipulation is not new; after all, viral RNA and DNA have abused human cells for their purposes since time immemorial. But if microRNAs from food change our gene expression, that gives the phrase “You are what you eat” a whole new meaning.
MicroRNAs in Food
If our food really contains microRNAs, that would mean that foods not only nourish us, but also deliver information—information that could switch our genes on and off. Some researchers, in another allusion to dark matter, referred to microRNAs in food as “dark nutrients” and claimed they play an “important role for human health.” Yes, plant microRNAs enter human cells and can change gene expression, but let’s take a breath first. Would microRNAs in food even survive cooking and digestion?
Some originally plant-based industrial products such as olive oil and beer appear to have lost their microRNAs during the production process. Do microRNAs get lost on the stove as well? In the past we thought genetic material was destroyed by cooking, but newer experiments show that some plant microRNAs can withstand heat. Some, such as miR-159 in broccoli, remain stable after cooking, while others, such as microRNA-319 in artichokes, are partially destroyed. And the levels of other microRNAs, such as those in cooked beans and brown rice, even increase after cooking—presumably because they are released into the cooking water. Mammalian and avian microRNAs in meat, dairy products, and eggs survive cooking and processing, as investigations of pork and poultry sausages, ham, salami, hard-boiled eggs, cheese, and pasteurized milk showed. There were hardly any changes in microRNA levels between raw and roasted beef; however, they would still have to survive the acid bath in the stomach.
Here, too, the scientific consensus held that microRNAs are destroyed during digestion, but when you immerse them in acidic gastric fluid, most plant microRNAs appear to survive at least six hours. In the small intestine, however, there are ribonucleases—enzymes that break down naked RNA. Do microRNAs survive this gauntlet? Perhaps they do not have to. A study in mice found that microRNAs from food are absorbed into the bloodstream already in the stomach. Or microRNAs move packaged in protective exosomes.
Vesicles from plants, similar to exosomes, are now known as “edible nanoparticles,” and they can be full of microRNAs. A pound of fruit can contain a gram of these little courier packages. This kind of packaging has been proposed as the solution to the question of microRNA bioavailability. It may be that microRNAs in this form are absorbed by the intestinal mucosa, repackaged into exosomes, and then handed off to the bloodstream. But the proof of the pudding is in the eating—if we eat microRNAs, do they actually show up in the bloodstream?
Unlike typical animal microRNAs, the tips of plant microRNAs are marked with a methyl group. This not only makes them more resistant to digestion, but also allows researchers to distinguish them from microRNAs already present and circulating in animals. If you feed mice cruciferous vegetables, then crucifer microRNAs reach their peak in the animals’ blood within six hours and can be detected in multiple organs. In the blood of pigs fed corn, corn microRNAs reach their peak level between six and twelve hours after consumption. Most plant microRNAs are transported in exosomes, which can also bring RNA into the brain. The crucifer microRNAs could be detected in the animals’ blood for more than 36 hours. And what about in humans?
Researchers found that 5 percent of all microRNAs detectable in the human body could come from plants. The first study on plant microRNAs in humans was published in 2012 and showed that rice microRNAs circulated consistently in the blood of Chinese consumers. Just as fish microRNAs circulate in fish-eating seals and plant microRNAs from feed plants and grasses circulate in cattle, most plant microRNAs in humans come from fruits and vegetables. Plant microRNAs were found throughout the human body, including the brain, breasts, kidneys, liver, and lungs, as well as in breast milk, amniotic fluid, and umbilical cord blood. Are these chance findings, or are the microRNAs from food harmful or beneficial to us?
MicroRNAs in Fruits and Vegetables
Hundreds of different microRNAs were found in the edible nanoparticles of common fruits and vegetables. In a basic research study, edible nanoparticles from grapes were fed to mice. The particles were taken up in the mice’s intestinal cells, changed their gene expression, and protected the mice from intestinal inflammation. Similar experiments with carrot, ginger, and grapefruit nanoparticles produced a whole range of beneficial regulatory effects, but how do we know that the microRNAs were actually responsible?
MicroRNAs are such simple molecules that we can synthesize them from scratch. So researchers made synthetic strawberry microRNA-156, rice microRNA-168, and cabbage microRNA-874 to isolate microRNA-specific effects. And in fact, these had anti-inflammatory effects on human cells. RNA extracts from blueberries, raspberries, and apple peels had a similar effect. To rule out the possibility that this was a generic RNA effect, an RNA extract from beef was also tested, and it did not relieve the inflammation.
One plant microRNA that circulates in humans is microRNA-156a. Its level was reduced in the blood and blood vessels of patients with cardiovascular disease, suggesting that it could have a protective effect. But where can microRNA-156a be found in concentrated form? In green vegetables. When a person eats a salad, their microRNA-156a rises significantly within an hour. Could lower microRNA-156a concentrations in cardiovascular patients be an indication of low consumption of green vegetables? To find out whether this is truly cause and effect, researchers exposed human arterial endothelial cells to pure (synthesized) microRNA-156a and showed that it targets a sticky protein called Junctional Adhesion Molecule-A, which promotes the attraction of inflammatory immune cells into the arterial wall to form atherosclerotic plaques. And indeed, increasing microRNA-156a reduced the adhesion of inflammatory cells to the arterial wall cells. That means the protection conferred by green vegetables against cardiovascular disease may be more than the effect of nitrate.