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Milk has evolved through mammalian history as a soup of complex molecules that provide nutrients, as well as developmental and immunological support to infants. Some of these complex molecules have been naturally selected for their abilities to deliver bioactive compounds in such a way that the infant body can make use of them. This involves, for example, the ability to bind ions with positive and negative charges, such as iron and calcium ions, respectively—and protecting delicate compounds from stomach acids so they can be absorbed through the intestinal wall. In short, some of the soup of complex molecules in milk are ready-made nano-scale delivery units that could be harnessed by science to carry modern medicines into the body to precise locations.

Researchers are already tapping milk’s chemistry for safe molecular mules to transport pharmaceuticals and dietary supplements into the human body. The main proteins they have to work with are of two categories: caseins and whey proteins. Caseins—which make up 79% of milk’s total protein [1]—are a family of proteins that, in a watery solution, naturally organize themselves into spheres, called micelles. For the young infant this means that caseins are able to provide lots of calcium in addition to the nutritionally essential amino acids of which they are composed [1]. The fine details of how exactly casein micelles interact with calcium are still debated, but it is thought that small casein micelles bind to colloidal lumps of calcium phosphate and then group together [2].

For an infant, caseins’ structures allows calcium phosphate to be released and then coagulate in the stomach. This is because caseins are rich in an amino acid called proline, which contributes to the interactions—or bonds—that hold together casein micelles, creating a relatively open structure that is accessible to stomach enzymes [1]. To the researcher looking for a safe way to target pharmaceuticals to the stomach, for example to treat gastric diseases, caseins offer a potentially very useful vehicle. Moreover, work to date shows that it is possible to bond nutraceuticals and synthetic drugs to casein proteins using various laboratory techniques, from heat-gelation to “polyelectrolyte ionic complexation” [1].

However, some of the most notable progress in this field has focused on the other main protein category in milk. Whey proteins, which comprise 19% of milk’s total protein and are a waste product of cheese-making, are mostly made of up the proteins β-lactoglobulin and α-lactalbumin [1]. It is the former of these that is currently exciting researchers. In some ways, β-lactoglobulin is a complementary tool in the drug delivery toolkit to caseins. It is stable to the acid environment of the stomach, and so has the potential to protect otherwise delicate molecules through that gauntlet of the gut, enabling them to be released later in the digestive process. Moreover, like caseins, there are options for chemically linking β-lactoglobulin to molecules that one might wish to carry into the body, and it can bind to hydrophobic molecules (that is, oil-loving, as opposed to water-loving chemical structures) [1].

The efforts of several groups of researchers have recently demonstrated the potential for taking advantage of β-lactoglobulin’s chemical characteristics. One group, based in China and South Korea, has made progress using it to enable the slow release of a component of green tea with anti-oxidant properties, called epigallocatechin gallate (EGCG) [3]. EGCG is normally given as a supplement encapsulated in chitosan, a long-chained carbohydrate molecule made from treating crustacean shells with a strong alkali. The reason for using chitosan is that it sticks to the walls of the intestine, allowing what’s left of the EGCG by that point in digestion to hang around in the intestines for longer. Only from there can EGCG be absorbed into the blood, and thereafter, as it circulates around the body, ward-off cancer and have other positive health effects that are linked to anti-oxidants.

The existing problem with delivering EGCG in this way is that chitosan does little to protect it from the ravishes of stomach acid. This is why Jin Liang of Anhui Agricultural University’s “State Key Laboratory of Tea Plant Biology and Utilization” in Hefei, China, and colleagues, decided to try coating it in both chitosan and β-lactoglobulin. They compared their double-layered approach with the usual chitosan coating in a series of laboratory experiments intended to allow them to evaluate changes to EGCG availability over the course of a mimicked digestive process.

They mimicked the stomach by adding pig pepsin and acid in a shaking water bath. Then they continued the shaking, increased the pH from 5.3 to a more neutral 6.8 by adding sodium hydroxide, and injected lipase, pancreatin, and bile extract from pig pancreas. Only after all of this physical and chemical processing would the chitosan-EGCG samples—and the chitosan-EGCG encased in β-lactoglobulin—be in the kind of state that they would be in after travelling through the small intestine. β-Lactoglobulin is degraded by pancreatic enzymes, so at this point it, too, would have been stripped away.

After analyzing samples drawn from all along the mimicked digestive process, the researchers concluded that pepsin-resistant outer layer of β-lactoglobulin helps chitosan-EGCG complexes reach the intestines relatively unscathed. As a result, the amount of EGCG left to be released in the intestines was higher. Furthermore, when they arrived in the intestines, the chitosan chains that had adhered to the β-lactoglobulin were more exposed—having had the β-lactoglobulin stripped off—than they otherwise would have been, which prolonged the length of time for EGCG absorption because the exposed chitosan proved even better at sticking to the intestinal wall.

The targeting of drug delivery that β-lactoglobulin makes possible is also being exploited by pharmaceutical researchers, Vilasinee Hirunpanich Sato, of Mahidol University in Bangkok, and Hitoshi Sato, of Showa University in Tokyo. Rather than EGCG, they are looking to deliver the immunosuppressive drug cyclosporin A to the intestines for absorption, using β-lactoglobulin as a gastric-juice shield. Currently, synthetic absorption enhancers are used to target cyclosporin A, and there is concern that these may have toxic side-effects.

So far, the pair’s experiments feeding rats have been promising. The results have shown that using β-lactoglobulin instead of synthetic alternatives provides gastrointestinal absorption of the drug, and faster absorption as well [4]. In a paper published in the International Journal of Drug Delivery, they propose that other researchers view their experiments as a proof of concept for deploying β-lactoglobulin to carry all kinds of orally-delivered lipophilic medicines (drugs that dissolve in oil, not water) to where the body can actually pick them up, and add them into the bloodstream. In short, their argument is that there is a huge opportunity to take the primary constituent of a cheese-making waste product and use it to make many medicines work better.

References

Human beings have internal clocks. Locked in a room with no source of daylight nor regularly scheduled stimulation, our bodies cycle automatically through periods of slightly longer than 24 hours, sleeping and waking more or less as if the sun were rising and falling over a horizon that we could see [1]. But we are not born this way. Instead, infants develop body clocks gradually. Researchers investigating this aspect of development have recently wondered how much human milk contributes to the process, in the knowledge that its levels of nutrients and hormones vary over the course of the day.

In a recent issue of the journal Pediatric Research, Jennifer Hahn-Holbrook, an assistant professor at the University of California, Merced, and her colleagues, point out that no published research has measured how daily variation in levels of cortisol circulating in the blood develops in infants who were breastfed, compared with those who received formula [2]. Cortisol is a hormone involved in regulating metabolism, and is generally high in the morning, when your body senses that it is time to wake up.

Indeed, studies into the potential benefits of breastfeeding over formula feeding often fail to note whether infants were fed at the breast, or with milk that was pumped and stored, potentially during a different portion of the day. And yet, as the authors of the review propose, infant body clocks may develop more rapidly among infants who consume milk that is laced with hormones appropriate to time of day that they feed. Hahn-Holbrook and her colleagues expect that these infants’ body clock development outpaces both formula-fed infants and infants who consume breast milk containing confused circadian signals.

Human milk changes over the course of the day in its levels of many different constituents. Concentrations of amino acids alter such that more activity-promoting amino acids are found in day milk than in night milk [2]. One example is tyrosine, an amino acid that is converted in some brain cells to a precursor of epinephrine and of the neurotransmitter, norepinephrine. Levels of some nucleotides also change. At night, infants fed at the breast receive more 5′AMP and 5’GMP than when they feed during the day. These nucleotides are thought to prompt the release of melatonin—a hormone that promotes sleep—and of gamma-aminobutyric acid (GABA), which does the same [3]. But it is two hormones in milk that are suspected of having the greatest influence in regulating the infant body clock, and in aiding its development: the aforementioned melatonin, and cortisol. This is where the bulk of the research in this burgeoning field has been focused.

Cortisol shows dramatic swings in its presence in human milk. One study, based on milk produced by 23 women, found more than three times the levels of cortisol in milk pumped between 4 a.m. and 10 a.m., compared with milk that the same women pumped between 4 p.m. and 10 p.m. [4]. Without research to actually test the hypothesis, this kind of finding is highly suggestive of impacts on infant behavior and development. This is because cortisol in the diet is known to easily pass through the intestinal wall into the blood and on, through the blood-brain barrier, into the brain. And because dosing rats in a laboratory with a synthetic version of cortisol, called dexamethasone, has been shown to reset the clock-following chemistry of their peripheral tissues. Specifically, in a series of experiments, Ueli Schibler and a team based at the University of Geneva, in Switzerland, showed that giving lab rats dexamethasone at times of day when their natural levels of cortisol were not high, changed the circadian gene expression patterns of liver, kidney and heart cells, effectively dialling these cells’ internal clock hands backwards or forwards by several hours [5].

As with cortisol, melatonin can cross through the walls of the intestine and make its way into many organs, including the brain. There is a study that directly compared the daily variation in a metabolite of melatonin in breastfed and formula-fed infants. It found, as anticipated, that the urine of breastfed infants contained more regular night-time increases in this metabolite—a biochemical trace of more rapidly developing sleep patterns in these infants [6]. Breastfed infants are known to have less fragmented sleep and higher sleep efficiency than formula-fed infants [7, 8], and this may be an important reason why.

As Hahn-Holbrook and her colleagues argue in their review of the field, this is an area of science ripe for policy-relevant research. Many of these experts’ suspicions about how human milk’s daily biochemical variations are likely to affect infant body clock development, if confirmed, would suggest basic and rather easy interventions. For example, labeling pumped milk with the approximate period of the day—night, morning, afternoon—and trying to match the pumping period to the feeding period, would be an easy way to avoid mixing the biochemical signals of alertness and sleepiness that human milk is thought to communicate to an infant body. For any parent who yearns for their infant to start sleeping through the night, this could help. But as Hahn-Holbrook’s review points out, these potential effects are off-the-radar of most of the medical profession.

References

Aging-related ailments can interfere with the daily life of the elderly. Older adults are at greater risk of diseases such as dementia or cardiovascular and orthopedic diseases. These diseases can contribute to functional disability—a decrease in physical, cognitive or emotional functioning that results from a health condition and adversely affects a person’s daily personal and social activities (1,2). Researchers have thus been looking for ways to decrease functional disability in the elderly.

Dairy intake has been reported to contribute to a lower risk of diseases such as dementia, cardiovascular disease, and orthopedic disease (3-7). A recent study by Dr. Sandra Iuliano at the University of Melbourne also found that dairy consumption was associated with favorable cholesterol ratios in older adults (8). These studies suggest that dairy intake may have some favorable effects in the elderly; however, little is known about the association between dairy intake and functional disability (9,10). “The literature is still quite mixed, not necessarily because there’s no effect, but because these kinds of studies are really hard to do,” says Iuliano.

In a new study, Dr. Yoshida and his colleagues at Kyushu University in Fukuoka, Japan examined the association between dairy intake and the development of functional disability in an elderly Japanese population (11). They found that higher dairy intake was associated with a lower risk of functional impairment, as well as with a lower risk for progression of functional disability. “The researchers saw a favorable association between function and dairy intake,” says Iuliano.

“The association seen in the study seems quite reasonable, as providing dairy protein in combination with exercise is likely to provide the ability to maintain function,” says Iuliano. “One take-home message of the study is that dairy intake may be a good avenue for protein,” she says. “But there could be other components involved that we don’t understand,” says Iuliano.

The results suggest that dairy products may have a protective influence on functional disability and its progression in the elderly. However, Iuliano notes that the new study is still quite exploratory. “It can generate thought, and from this thought we can generate better trials,” she says.

In the new study, the researchers analyzed data from an ongoing population-based study that is designed to evaluate the risk factors for lifestyle-related diseases (12). Eight hundred fifty-nine elderly Japanese residents of Hisayama without functional disability were followed up for seven years. The residents’ dairy intake was evaluated using a diet history questionnaire, and the researchers assessed their functional disability at baseline and at follow up.

The researchers found that the risk of developing functional impairments, especially impairments in intellectual activity and social roles, was significantly lower in participants who consumed dairy food compared with those who did not. These kinds of impairments are thought to develop at early stages of functional disability (13, 14). The risk of more severe functional disability also decreased significantly with higher dairy intake, even after accounting for various confounding factors.

Three major diseases—dementia, stroke, and orthopedic disease—were responsible for most of the functional disability in the study population, and the researchers suggest that dairy intake may protect against functional disability by helping to prevent these diseases (15).

The study also suggests that the protein component of dairy might be playing a role in decreasing the risk of functional disability. “Protein could be a key driver of the effect, as when the researchers adjusted for protein a lot of the association went away,” says Iuliano. There was also a strong positive association between dairy and total protein intake in the study participants. “So dairy was the main source of protein,” she says.

It’s unclear how generalizable the study findings are to other ethnic populations with different lifestyles. “We need to look at the dairy intake in the context of the country,” says Iuliano. She suggests the need for additional long-term studies. “The ideal would be to follow a large group of people over time, with half getting extra dairy food and the other half continuing with their normal diet,” says Iuliano.

Iuliano also suggests that researchers could monitor ongoing trials more frequently, maybe by assessing dietary intake once a year so that they can look for trends. “If we’re monitoring participants over time, we can see when functions decline and look at whether there were changes in their dietary pattern then,” she says.

The study concludes that higher dairy intake is associated with a lower risk of functional disability and its progression in the elderly, and suggests that dairy products could have a protective effect against functional disability. But more studies will be needed to confirm these results and examine the mechanisms by which dairy could be having these effects.

“The beauty of dairy foods is that there are lots of things we still don’t know,” says Iuliano. “There may be interactions during fermentation, or there may be components that in isolation do one thing and when they’re combined do another thing,” she says. “That’s what makes it so complex and fascinating to study.”

References

It’s a tough gig being a cow. Productivity expectations for meat and milk are high, and at the same time, the cow gets a bad rap for belching a potent greenhouse gas, methane, which is a by-product of its digestion. Some people say it’s like driving a car very hard on a winding mountain road and then complaining about the car’s increased exhaust gas emissions. Reducing emissions and fuel consumption while maintaining performance is the golden ambition of car manufacturers. A similar goal is also true for the cow. People in many government agricultural agencies and the Food and Agriculture Organization (FAO) want the beef and dairy industries to use more productive cattle emitting less methane and using less feed i.e., increasing industry production efficiency while decreasing its environmental footprint [1-4]. It’s a tall order seemingly resisted by the realities of cow biology, however recent ground-breaking research may have opened new opportunities to meet these ambitious aims [5].

Methane emission from belching cows may be modifiable by changing a specific population of microbes in the cow’s rumen, according to a large group of international investigators belonging to the aptly named RuminOmics research project [5]. The rumen is a large compartment in the cow’s stomach where most of the digestion of forage occurs. In particular, the investigators suggested that livestock could be selectively bred to contain specific rumen microbes producing lower levels of methane. Their research is timely and it potentially has important implications for decreasing methane levels in the atmosphere.

Methane Is a By-Product of Digestion

The digestive system of ruminant animals is amazing. Ruminants are hoofed mammals like cows, sheep, and goats that continually rechew their ingested feed. These animals efficiently convert pasture into nutrient-dense meat and milk for human consumption. The survival of humans over the ages has been tightly intertwined with the hunting and domestication of ruminant species, which provided humans with a reliable, mobile, and highly nutritious food source [6]. Today, the human population still relies on ruminants as a primary food source. Importantly for humans, ruminant animals can often prosper on plant fiber obtained from pasture growing on semi-arid land that is unsuitable for other agricultural purposes. The cellulose in plant fiber contains a lot of energy, but ruminants need help from microbes to tap into it.

The cow stomach has four large compartments, the reticulum, omasum, abomasum, and rumen. The rumen is where most of the digestive action occurs. Its volume is large, about 150 litres (40 U.S. gallons), and each milliliter of rumen fluid contains billions of microbes [7]. Thus, the cow rumen holds an astonishing number of microbes called the rumen microbiome. The complex microbial ecosystem in the rumen efficiently digests plant fiber in the absence of oxygen to produce small molecules that provide the cow with essential nutrients and energy.

The microbial ecosystem in the rumen contains a huge diversity of microbes including, in order of abundance, bacteria, archaea, ciliated protozoa (moving single cells that have a nucleus like animal cells), and fungi [8]. The archaea are single-celled organisms with similarities to bacteria, but they also have important metabolic differences. The bacteria, protozoa, and fungi work together using a fermentation process to degrade ingested forage and convert it into chemical energy for their use. These microbes also produce small fatty acids as side products of the fermentation, which are absorbed by the cow for use as its primary energy source.

The microbial fermentation process in the rumen also produces carbon dioxide and hydrogen gas, which are used by some of the archaea (hydrogen is their energy source). An important by-product of archaea metabolism is methane, which the cow releases into the atmosphere by belching. The emission of methane also represents wastage of up to 10% of the total energy in the cow’s ingested feed [2]. Less energy means less meat and milk. Thus, decreasing methane emission from the cow could be good for the atmosphere and could also improve dairy cow productivity. The key to decreasing methane emission from the cow is to decrease some of the archaea microbes in the rumen.

Methane Metrics

The FAO estimates that there are about 3 billion domesticated ruminants in the world [1] that release approximately 100 million tons of methane into the air each year [2, 9]. Most of the methane emission comes from about 1.5 billion beef and dairy cattle [2]. Over the next 30 years, the FAO predicts a doubling of global demand for meat and milk as the human population heads towards ten billion people [9, 10]. The extra food demand will require increased numbers of ruminants that further increase the level of methane in the atmosphere [9]. The numbers are all big, getting larger, and demanding attention. So, why is there such a fuss about methane?

Methane is approximately 30 times more potent than carbon dioxide as a greenhouse gas [2, 9]. On the brighter side, the lifetime of newly emitted methane in the atmosphere is only about ten years while the carbon dioxide lifetime is 100 years [9]. A decrease in methane emission from ruminant animals can have relatively rapid beneficial effects on the load of greenhouse gases in the atmosphere. Consequently, strategies for methane abatement from ruminant livestock industries are a high priority of many government agencies [1, 2, 11-13].

Deep Exploration of Rumen Microbes

A large group of scientists from eight countries recently published their new and innovative insights into the complex microbial communities present in the cow rumen [5]. The first author on the publication is John Wallace from the University of Aberdeen. Their research, published in Science Advances, highlighted international cooperation at its best. The investigators’ aims were first to determine whether cow genetics regulates the different populations of microbes in the rumen, and secondly to identify the specific microbial populations in the rumen that affect cow productivity traits and methane emission. This milestone investigation used more than 1,000 Holstein and Nordic Red dairy cattle from the U.K., Italy, Sweden, and Finland.

Wallace and colleagues made detailed measurements on the cows for milk output and composition, feed intake and digestibility, blood plasma components, and methane and carbon dioxide emissions [5]. The different types of microbes in the rumen (the rumen microbiome) and the genetics of the cows were also determined. The investigators showed that the cow population contained substantial genetic variation, which potentially could contribute to production trait differences between individual cows. Wallace and colleagues generated an enormous amount of data. The investigators first identified a core microbiome consisting of about 500 microbial species. The core microbiome is a population of microbes that is present in at least half of the cattle population. It only represented a tiny percentage of the total diversity of microbial species present in the rumen, yet the core microbiome was highly abundant in terms of microbial numbers. Wallace and colleagues used information from other published studies to conclude that this core microbiome is characteristic of most other ruminants. Hence, many of their research conclusions may also apply to other ruminants, especially beef cattle.

The investigators discovered mathematical correlations between a subset of the microbial species in the core microbiome and genetic differences in the cow population [5]. This was a remarkable and important discovery as it inferred that cow genetics somehow determined the presence of particular species of microbes in the core microbiome of the rumen. Hence, cow genetics potentially indirectly influenced the microbial metabolic activity of rumen. How this occurs is still a mystery. More specifically, Wallace and colleagues pointed out that some microbial components of the “inherited” core microbiome were already implicated in differences in methane emission in individual cows.

Wallace and colleagues used advanced computational analyses to determine whether there were associations between the microbial composition of the core microbiome and the various dairy production trait measurements [5]. They struck gold and discovered that components of the core microbiome were linked to many of the cow production measurements including milk output and milk composition. Some of the links were surprisingly strong while others were weaker. The investigators had linked together cow genetics, various cow production traits, and components of the core microbiome in a triangle of interdependent new knowledge. In particular, the investigators identified a “heritable” component of the core microbiome that was linked to methane emission. There have been hints from other researchers that cow genetics influences methane emission [14-16]. However, the study performed by Wallace and colleagues was the first comprehensive and detailed analysis. The race must be now on by scientists to discover the mysterious cow genes that affect the methane-generating microbes in the core microbiome in the rumen.

Implications

Wallace and colleagues’ provocative conclusion is that it may be possible to breed cattle with optimal rumen microbial populations that potentially maintain the high production performance of dairy cows but emit decreased quantities of methane [5]. This strategy would generate a permanent decrease in methane emission from the selectively bred cattle. The investigators also noted that a more rapid but non-permanent decrease in methane emission could also be achieved by early-life inoculation of the calf rumen with key populations of the core microbiome. Wallace and colleagues suggest that the broad application of these collective approaches in all ruminant livestock animals “should result in a more efficient and environmentally friendly ruminant livestock industry.” Cows are about to go green.

References