Sunday, September 20, 2015

Burgers on breasts – what is really going on here?

Recently, a series of advertisements came into the public consequence. These images depict women breastfeeding, with one breast painted to look like a hamburger, doughnut, or can of soda. The message is “Your child is what you eat”, suggesting that the mother’s diet is directly related to the construction of the baby – that is, eat a burger, build a baby from burgers.

Before we get into a discussion of the factual accuracy of the statement, it is worth taking a moment to think about the ads themselves. The ads were most likely produced by the advertising company PAIM, and the logo depicted is that of the SPRS – the Pediatric Society of Rio Grande (Brazil). SPRS is sponsored by Nestle, and there is some evidence that PAIM also holds the Nestle ad campaign for Brazil. But here’s my big question: why are the ads in English? The national language of Brazil is Portuguese, and as of 2011, the best guess for English fluency among educated Brazilians was 8-11% (Glickhouse, 2012). Your milk what you eat should be “Seu leite é o que você come” (if Google translate is accurate). 

Although the images contain the logo for SPRS and a link to the website, the ads are not on the SPRS website. The SPRS website is also in Portuguese. Moreover, the food images just don’t quite match up to typical Brazilian cuisine – burgers commonly have eggs or corn for example, and doughnuts with holes in the middle are not that common in Brazil. So – just who were those ads designed for?
Leaving aside this very big question, the ads have generated considerable international attention and debate. Many mothers have expressed concern about the message: your baby is what you eat. But is it true? Well, the best answer is “somewhat – but not really”, at least as a far as breast milk is concerned.

Breast milk is produced by the mothers’ body (for review, see my prior post here). Generally, the proteins and sugars in the milk are largely independent of what the mother eats. This reflects the synthesis of protein and sugar by specialized cells, called lactocytes, in the breast. Synthesis occurs from glucose, amino acids, and similar. Very few dietary or lifestyle factors appears to influence milk protein, especially milk proteins that are not immunoproteins. 

Total milk fat, at least for humans, is largely independent of the mother’s diet (Villalpando and Del Prado, 1999). However, the fatty acid composition of the milk fat is strongly correlated with the maternal diet.

Probably the best known example is that of DHA. Most pre- and post-natal vitamins contain fish oils or DHA. DHA is a long chain, polyunsaturated fatty acid that cannot be produced by the mother’s body and must be derived from her diet. Ergo, milk DHA is very sensitive to the mother’s diet – more dietary DHA, more DHA in milk. Figure 1 shows an example from our work in the Philippines (Quinn et al., 2012), where we found a dose association between meals of a local fish (bodboron) and milk DHA. If you haven’t heard of bodboron, don’t be surprised – in the US, it is commonly fed to salmon rather than consumed. Working with mothers in Denmark, Lauritzen et al., (2002) also showed a positive association between fish consumption (in this case salmon) and milk DHA. Fish oil consumption has long been recognized as a source of milk DHA (Harris et al., 1984). It is also understood that fatty acids – especially long chain fatty acids like DHA – accumulate in body fat where they can be utilized later. 
Figure 1: Fish consumption and milk DHA content in a sample of Filipino mothers living in Cebu, Philippines (Quinn et al., 2012).

What if a mother is consuming a diet that is very low in fat? There is no substantial evidence that her total milk fat will be lower (Villalpando and Del Prado 1999). This is another one of those instances where mothers’ bodies are amazing – her body will produce certain fatty acids from glucose derived from the carbohydrates in her diet. Humans cannot produce long chain fatty acids like DHA – but the breast and liver can produce medium chain fatty acids – fatty acids with less than 14-16 carbons (Figure 2).  These medium chain fatty acids are routinely incorporated into milk fat, and, when dietary and existing fat stores are insufficient to meet demand, the mother’s body will produce more medium chain fatty acids to maintain milk fat (Rudolph et al., 2007). Thus, total fat is maintained, even if fatty acid composition is altered.

Figure 2: Comparison of two fatty acids - TOP: lauric acid, which has a carbon chain length of 12 and can be produced by the mammary gland and liver, and BOTTOM: DHA, which cannot be produced by the body.

Another concern would be man-made food products, like trans-fatty acids or artificial sweetners (up next!). These have been found in human milk; and there is general concern about the levels of trans-fatty acids in human milk. Product labeling has resulted in lower dietary intakes and a reduction in milk trans-fatty acids in Canadian mothers (Ratnayake et al., 2014); a recent analysis of some 2327 food products in Brazil found that more than half contained trans-fatty acids. The worst offenders were cookies, so if this was really an ad about trans-fatty acids, a cookie would be the appropriate image.

What does it come back to? These advertisements are making the same fictional point we have discussed here before with medical devices. You are a threat to your baby. Your body can’t be trusted. You can’t be trusted to eat right for your baby and thank heavens there is formula so you can’t harm your baby with your hamburgers.  These ads are deliberate, and the target audience is very much open for debate.

Glickhouse R. (2012) Lost in Translation. Christian Science Monitor.

Lauritzen L, Jørgensen MH, Hansen HS, Michaelsen KF. (2002) Fluctuations in human milk long-chain PUFA levels in relation to dietary fish intake. Lipids 37(3):237-44.

Quinn EA, Kuzawa CW. (2012) A dose-response relationship between fish consumption and human milk DHA content among Filipino women in Cebu City, Philippines. Acta Paediatr 101(10):e439-45. 

Ratnayake WN, Swist E, Zoka R, Gagnon C, Lillycrop W, Pantazapoulos P. (2014) Mandatory trans fat labeling regulations and nationwide product reformulations to reduce trans fatty acid content in foods contributed to lowered concentrations of trans fat in Canadian women's breast milk samples collected in 2009-2011. Am J Clin Nutr 100(4):1036-40.

Rudolph MC, Neville MC, Anderson SM. (2007) Lipid synthesis in lactation: diet and the fatty acid switch. J Mammary Gland Biol Neoplasia 12(4):269-81.

Villalpando S, del Prado M. (1999) Interrelation among dietary energy and fat intakes, maternal body fatness, and milk total lipid in humans. J Mammary Gland Biol Neoplasia 4(3):285-95.

Saturday, May 16, 2015

Milk responds: Changes to milk immune factors with infant (or maternal) infection

FOREWARD. As many of you know, my recent research has been looking at milk composition, infant growth, and maternal health in a population of ethnic Tibetans living in the Himalayas of Nepal. The communities I work with were within 50 miles of the epicenter of the earthquake on April 25.  While the loss of life in these communities was minimal (thankfully), there was considerable destruction of homes, clinics, schools, and infrastructure (water, latrines). With the upcoming monsoon season, there is considerable need for safe drinking water, food storage, medical care, and safe homes. Several NGOs with long standing relationships with the communities are currently fundraising for relief and rebuilding efforts. Please consider donating to these organizations if you can afford to do so (NepalSEEDS; Tsum-Nubri Relief Center). We are still committed to these communities, and will continue to support infrastructure and research to promote maternal and child health.

In the last blog post – January – I discussed the idea of immunological memory in milk, particularly the well described association between maternal exposure to pathogenic bacteria in early life and the immunological memory of those bacteria, by specific forms of secretory Immunoglobulin-A (sIgA), many years later. Milk is incredibly dynamic, and this is certainly true for the immune factors in milk. Three recent papers have investigated this responsiveness in several samples, using a variety of immune factors to measure immune activation in milk.

Breakey et al., (2015) have articulated this as a model of two systems within the mammary gland – a protective paradigm, where some immune factors in milk are always protecting against infection; and the responsive paradigm, where active infection will increase the concentrations of immune factors in milk. Of the hundreds of known (and many unknown!) immune factors in milk, some will be generally protective, and others will be responsive (Brandtzaeg 2010). A few, including secretory IgA, will be both.  

Breakey et al., (2015) investigated the responsiveness of immune factors in milk to current infection using two biomarkers – sIgA and lactoferrin – in a sample of 29 Toba mother-infant dyads followed longitudinally. Both of these biomarkers have come up before (for reviews: sIgA; lactoferrin).The Toba are indigenous population from Argentina (Figure 1); previous generations have subsisted as foragers, but more recently the population has become increasingly concentrated in peri-urban areas, often in informal settlements lacking access to sanitation and water facilities. 
Figure 1: Location of the Toba. Image from wikicommons, author Nazareno98; produced in 2008.

Milk samples and interviews were collected monthly, allowing for the researchers to investigate milk composition before, during, and after an infection in the infant. Infant infections during the preceding month were collected during monthly interviews; all infants in the study had at least one illness over the course of the longitudinal study. Mothers did not report frequent illnesses, although this may have been underreporting. 

In this sample, infants receiving milk with higher sIgA were less likely to be ill, while infants receiving milk with more lactoferrin were more likely to be ill. Although causation cannot be certain, the authors hypothesize that lactoferrin content of milk increases during an infection (responsive) while sIgA levels are more generally protective. 

The study with the Toba follows two earlier studies of immune responsiveness in milk, both done in WEIRD populations. The earliest, by Riskin et al., (2012) remains one of my favorite papers. In this study, Riskin et al., recruited 51 mother-infant dyads, younger than 3 months, from Haifa, Israel into the study. 31 mother-infant pairs were hospitalized for fever at the time of recruitment, with an additional 20 pairs serving as healthy controls. Milk samples were collected from the mothers while the infants were hospitalized, and then seven days later; samples from controls were collected at one week intervals. Milk samples were analyzed for immune cells (lymphocytes, neutrophils, macrophages, CD45+), sIgA, lactoferrin, TNF-alpha, and IL-10 (Figure 2). 

Figure 2: Important cells in the immune system. Not all are found in milk. Image credit:
For the purposes of analyses, the participants were grouped into 3 categories: controls (healthy mom, healthy baby; n=20), all sick (all infants sick, moms sick or not; n=31), and sick infant (only baby sick; n=20). For the control group, there were no changes in the immune factors measured in milk from time 1 to time 2. However, for the sick group, there were significant declines in CD45+ cells, lymphocytes, neutrophils, macrophages, IL-10, and TNF-alpha. Lactoferrin and sIgA also declined, but the differences were minor. It does not appear that the associations were simply responding to maternal infection either. In the 20 mothers of sick infants who were not ill themselves, milk cd14+ cells, neutrophils, and macrophages also showed a significant decline from the original to the after measure. All other immune factors also showed declines, but again these were relatively minor. 

In an additional study of 21 mother-infant pairs, Hassiotou et al., (2013) reported increased leukocytes, and sIgA in the milk of mothers with infections compared to earlier and later samples from the same mothers collected as part of a longitudinal study design. While both maternal and infant infection increased leukocytes and sIgA in milk, this was most pronounced for mothers with breast infections. 

One of the leading hypotheses for how maternal physiology may respond to infection in the infant is through oral contact. Saliva from the infant’s mouth may enter the breast, carrying the pathogens responsible for the infection. This would encourage a localized immune response to the pathogen in the mammary gland itself (Hassiotou et al., 2013), although Riskin et al., (2012) also propose a model of subclinical infection in the mothers.

The capacity for milk to balance between innate and adaptive immune responses is incredibly important, especially for infants living in highly pathogenic, low resource environments such as the Toba, or my own participants from Nubri (more on this to come). Certainly, having a milk to gut superhighway for immune factors should be incredibly important in promoting gut integrity, decreasing infant illness, protecting against growth faltering, and promoting infant survival. Infant – or maternal – illness becomes then not a reason to stop nursing, but a reason to nurse more.

Brandtzaeg P. (2010) The mucosal immune system and its integration with the mammary glands. J Pediatr. 156(2 Suppl):S8-15

Breakey AA, Hinde K, Valeggia CR, Sinofsky A, Ellison PT. (2015) Illness in breastfeeding infants relates to concentration of lactoferrin and secretory Immunoglobulin A in mother's milk. Evol Med Public Health. 2015(1):21-31. doi: 10.1093/emph/eov002.

Hassiotou F, Hepworth AR, Metzger P, Tat Lai C, Trengove N, Hartmann PE, Filgueira L. (2013) Maternal and infant infections stimulate a rapid leukocyte response in breastmilk. Clin Transl Immunology. 2(4):e3. doi: 10.1038/cti.2013.1
Riskin A, Almog M, Peri R, Halasz K, Srugo I, Kessel A. (2012) Changes in immunomodulatory constituents of human milk in response to active infection in the nursing infant. Pediatr Res. 71(2):220-5. doi: 10.1038/pr.2011.34.

Saturday, January 3, 2015

Milk remembers: Immune factors in milk “remember” childhood environments

It is well established that with very few exceptions, human milk is the preferred first food for infants. While the benefits of breastfeeding/receiving human milk are considerable and influence the development of multiple systems in the infant, perhaps the best known benefits of human milk are its immunoprotective properties. Worldwide, breastfeeding is associated with reduced risk of infectious diseases in infants, and these protections persist even in highly hygienic conditions such as the United States (Bartick & Reinhold 2010). Many immune factors are found in human milk, including immune cells, cytokines that regulate immune responses, and secretory Immunoglobulin-A (sIgA), perhaps the most common immunoprotein in human milk. It is well established that there is considerable variation in the immune factors in milk between individual mothers and between populations. It is also known that many of the immune factors in milk are highly responsive, changing in response to active infection of either the mother or infant (blog post on this topic coming next month). 

It has been traditionally held that the differences in immune factors in milk, especially sIgA, were reflecting the pathogenicity of the environment. The higher levels of sIgA found in the milk of women in developing countries was thought to be a proximate response to pathogen exposure in the immediate environment. However, some old – and some new – research suggests that the associations may be much more interesting. What if the past environment was just as important as the current environment in influencing sIgA and other immune factors in milk?

To study this, Nathavitharahna et al., (1994) decided to compare sIgA in the milk of women from three groups (sample size): women born in and currently living in Sri Lanka (n=64), women who had immigrated to England from Sri Lanka or other nearby countries in South Asia (n=20), and women born in and currently living in England (n=75).  Pathogen exposure for these groups broke down as follows: Sri Lankan women – high early life, high present; Immigrant women – high early life, low present; and British women – low early life, low present.
Surprisingly, as shown in Figure 1, there were no differences in the total amount of sIgA in the mean amounts of sIgA for each group – and within each group, milk sIgA ranged from 0.2 g/L to 19.1g/L! 

Figure 1: Comparison of total sIgA content in the milk of the three groups of mothers.
Mothers from England actually had more sIgA than mothers from Sri Lanka! However, total milk sIgA is only half the story. The researchers went on to look at specific sIgA antibodies to Escherichia coli (E. coli). They focused on 14 strands of E. coli commonly associated with moderate to severe diarrheal illness. Figure 2 is borrowed from the paper. Mothers living in Sri Lanka had the highest amount of E. coli specific sIgA in their milk. However, E. coli specific sIgA was much higher in the milk of immigrant mothers compared to their British neighbors. 
Figure 2: The amount of sIgA specific to each form of E. coli found in human milk for the three groups. Both horizontal and vertical axes are the same for each graph - white British women have much less E. coli specific sIgA in their milk than immigrant women or Sri Lankan women.

How can we interpret these findings? Nathavitharana et al., briefly considered that the pathogens may be maintained in the community, but further study demonstrated this was unlikely. Instead, the most logical explanation is that the sIgA in milk was the product of milk immunological “memory”. The sIgA in milk is produced by a type of immune cell called a B cell. These cells migrate to the mammary gland, often from the GI tract during last gestation/the onset of milk production. B cells include a special class of B cells, called memory cells. Memory B cells maintain “memories” of prior infections, allowing for a rapid antibody-mediated immune response should re-exposure occur. Memory B cells, it seems, were recording the mother’s own exposure history, migrating to the mammary gland, and providing infants with protection against the pathogenic E. coli experienced by their mother. Already having sIgA antibodies against common – and severe - pathogens may provide infants with increased capacity to resist or limit the severity of infection by these pathogens.

Six years later, another study, using a similar study design, provided further evidence for an immunological memory in milk. Holmlund et al., (2010) looked at three groups – women born and currently living in Mali (Africa), women who had migrated from Africa to Sweden (multiple countries represented), and women born and living in Sweden and analyzed their milk for several cytokines involved in immune function and sIgA. There were roughly 30 women in group. Holmlund and colleagues found few differences in the cytokines of the milks with two exceptions – Transforming Growth Factor beta ( forms 1 & 2). Women from Mali had the highest concentrations of each, with immigrant women having intermediate levels and Swedish women the lowest levels. Here’s the cool part – TGF-B2 is part of the signally cascade for sIgA, and sure enough – there was a significant association between TGF-B2 in the milk and sIgA. However, this association was only significant in the women living in Mali – but it was really, really close in the immigrant women.  

What does this all mean? It’s evidence for a complex immunological memory in milk. While this may not be important in highly hygienic environments such as the United States, it certainly suggests that there may be adaptive features in milk that record pathogen exposures during early life and provide a “dictionary” of potential infections to the infant. These highly specific forms of sIgA antibodies in human milk may allow for a more rapid immunological response by the infant. Milk “memories” therefore, would serve to protect the infant. How long these memories are retained is another question, and it seems likely that there will be rapid drift in the types of sIgA antibodies reflecting novel exposures by the infants. 

References - links included to open access papers

Bartick M, Reinhold A. 2010. The burden of suboptimalbreastfeeding in the United States: a pediatric cost analysis. Pediatrics 125(5):e1048-56. doi: 10.1542/peds.2009-1616.

Holmlund U, Amoudruz P, Johansson MA, Haileselassie Y, Ongoiba A, Kayentao K, Traoré B, Doumbo S, Schollin J, Doumbo O, Montgomery SM, Sverremark-Ekström E. 2010. Maternal country of origin, breast milkcharacteristics and potential influences on immunity in offspring. Clin Exp Immunol. 162(3):500-9. doi: 10.1111/j.1365-2249.2010.04275.x.

Nathavitharana KA, Catty D, and McNeish AS. 1994. IgA antibodies in human milk: epidemiological markers of previous infections? Archives of Disease in Childhood 71(F192-197).