Human Milk Oligosaccharides Alter Immune Cell Populations in Pigs

  • A new study examines the effects of human milk oligosaccharides (HMOs) and prebiotic oligosaccharides on immune cell populations in uninfected and rotavirus-infected piglets.
  • The study found that dietary HMOs altered systemic and gastrointestinal immune cell populations in piglets, and had a greater effect on immune cells than prebiotic oligosaccharides.
  • Dietary HMOs were previously shown to have some effects on rotavirus infection susceptibility, and the study suggests that HMO-associated changes to immune cell populations may mediate some of these effects.

 

Rotavirus is a major viral pathogen, and rotavirus-associated diarrhea is prevalent in many developing countries [1-4]. Interestingly, breastfed infants have a lower incidence of rotavirus infection than formula-fed infants, suggesting that certain components of human milk may have protective effects against this virus [5].

“Human milk is best, you’re going to have the best protection if you’re able to feed your baby human milk,” says Professor Sarah Comstock of Michigan State University. “We’re always interested in how we can improve the immune responses of formula-fed infants to make them more like those of human-milk fed infants,” she says.

Researchers have investigated whether sugars found in human milk, called human milk oligosaccharides (HMOs), might be responsible for some of the protective effects against rotavirus. HMOs are absent from most infant formulas and have antimicrobial and immunomodulatory actions in vitro [6-8].

In a new study conducted by Comstock and Professor Sharon Donovan of the University of Illinois, the researchers measured the effects of HMOs on immune cell populations from uninfected and rotavirus-infected pigs [9]. They also compared the effects of HMOs with those of prebiotic oligosaccharides. “We wanted that comparison group in there because prebiotics were already being added to infant formula, and it doesn’t make much sense to spend a lot of effort and money adding HMOs to formula if prebiotics can be just as effective,” says Comstock.

The researchers found that dietary HMOs altered systemic and gastrointestinal immune cell populations in pigs, and had a greater effect on immune cells than did prebiotics. The findings suggest that the HMO-associated change in immune cell populations may mediate protective effects of HMOs on rotavirus infection. “It’s just more evidence that breastfeeding really is good for your baby,” says Comstock.

Researchers have long been interested in studying the effects of HMOs on the immune system, but it’s only recently that they have been able to cost-effectively conduct such studies. “Up until the mid-2000s, even though there was an interest in the human milk oligosaccharides, it was really hard to cost-effectively synthesize them or to get access to enough HMOs to do these types of experiments,” says Comstock. “It finally seemed feasible to do this type of study, to actually feed a significant amount of human milk oligosaccharides to piglets and look at how it affected their response to infection,” she says.

Comstock and Donovan decided to test the effects of HMOs on rotavirus infection in a piglet model, whose immune and gastrointestinal systems share many similarities with those of humans. “When you combine the immune homology with this gastrointestinal homology, and you’re looking at a gastrointestinal virus, I think it’s just really the best system,” says Comstock. “The payoff in terms of using this type of model, which is so much closer to a human than using a rodent model, is huge,” she says. “I think we’ll begin to see more work in humans and I think you’ll see that the piglet research really is an important driver of what we might expect in those human studies,” says Comstock.

The new study builds on several previous studies by Comstock and Donovan looking at the interaction of HMOs, rotavirus and the immune system [7, 8, 10]. “We knew in vitro that HMOs were able to inhibit rotavirus infectivity,” says Comstock. “We had a lot of evidence that there were direct interactions of HMOs with rotavirus, which were preventing rotavirus from getting access to the intestinal epithelium,” she says. “Then we had evidence that feeding HMOs would definitely affect the types of microbes that were living in the intestine, and we additionally had evidence that HMOs could directly affect immune cells ex vivo,” says Comstock. “We wanted to put this all together and ask, are these effects happening in vivo,” she says.

In the new study, Comstock and her colleagues compared the immune cell populations of uninfected and rotavirus-infected pigs fed either a control formula, a mix of HMOs consisting of 2’-fucosyllactose, lacto-N-neotetraose, 6’-sialyllactose, 3’-sialyllactose, and free sialic acid, or prebiotics consisting of short-chain galactooligosaccharides and long-chain fructooligosaccharides.

Both infected and uninfected HMO-fed pigs had increased peripheral blood mononuclear cell natural killer cells and mesenteric lymph node memory effector T cells compared with pigs fed formula. The researchers also found that dietary prebiotics induced intermediate increases in immune cell populations compared with dietary HMOs. Prebiotic oligosaccharides may be less effective than HMOs because of differences in their structures or in the intestinal bacteria they affect.

The researchers concluded that dietary HMOs were more effective than prebiotics in altering systemic and gastrointestinal immune cells in pigs. Comstock suggests that the HMO-associated changes to immune cell populations may mediate the effects of dietary HMOs on rotavirus infection susceptibility.

Comstock notes that the current study only looked at the effects of 5 HMO structures out of the more than 200 HMOs identified so far. “Even from this limited complement of HMO structures, we’re able to get increased immune responsiveness, so imagine what it might be like if you were able to include a full complement of these 200-plus structures,” she says.

“Obviously it would be nice if we could get all 200 into infant formula; the more we can make infant formula look structurally like human milk, I think the better off those babies’ immune systems will be,” says Comstock. “Unfortunately, a lot of those are even more expensive or impossible to synthesize or extract,” she says.

Follow-up studies could look at the immune effects of HMOs in more detail, including analyzing their effects at different post-infection time points. “One limitation of this study is that we had to pick one time point to look at these effects,” says Comstock. “That does limit our knowledge in terms of the full complement of effects that HMOs can have on the immune response,” she says.

The researchers plan to further analyze their data to try to better understand how the HMOs affect rotavirus-infected piglets. Understanding the in vivo effects of HMOs could help researchers design improved infant formulas and find ways to protect against or treat rotavirus infection. “We do plan to do more detailed and sophisticated modeling of our datasets to see if we can gain any knowledge or generate any hypotheses about how the bacterial changes and the immune system changes are working together to resolve the infection and alter the clinical response,” says Comstock. “The question is, can we model those interactions to understand more fully what’s going on to reduce the length of diarrhea in the rotavirus-challenged piglets,” she says.

Future studies could also try to decipher the mechanism by which HMOs influence immune cell populations. “It would be great to really understand the mechanisms of what is happening,” says Comstock. “We don’t know if these HMOs are directly interacting with immune cell receptors and directly triggering these changes in the immune cells, or if through interactions with bacteria or viruses in the intestine they’re modulating the extent of the immune response that needs to happen,” she says.

“So there’s still a black box between feeding of the HMOs and the clinical response, but this study gives us some clues about what might be going on” says Comstock.

 

1. Bucardo F., Nordgren J. Impact of vaccination on the molecular epidemiology and evolution of group A rotaviruses in Latin America and factors affecting vaccine efficacy. Infect Genet Evol. 2015 Aug;34:106-13.

2. Tate J.E., Parashar U.D. Rotavirus vaccines in routine use. Clin Infect Dis. 2014 Nov 1;59(9):1291-301.

3. Bar-Zeev N., Kapanda L., Tate J.E., Jere K.C., Iturriza-Gomara M., Nakagomi O., Mwansambo C., Costello A., Parashar U.D., Heyderman R.S., French N., Cunliffe N.A.; VacSurv Consortium. Effectiveness of a monovalent rotavirus vaccine in infants in Malawi after programmatic roll-out: an observational and case-control study. Lancet Infect Dis. 2015 Apr;15(4):422-8.

4. Liu X., Meng L., Li J., Liu X., Bai Y., Yu D., Ren X., Liu H., Shen X., Wang P., Hu X., Wei K., Pei H., Kang Q. Etiological epidemiology of viral diarrhea on the basis of sentinel surveillance in children younger than 5 years in Gansu, northwest China, 2009-2013. J Med Virol. 2015 Dec;87(12):2048-53.

5. Plenge-Bönig A., Soto-Ramírez N., Karmaus W., Petersen G., Davis S., Forster J. Breastfeeding protects against acute gastroenteritis due to rotavirus in infants. Eur J Pediatr. 2010 Dec;169(12):1471-6.

6. Kunz C., Rudloff S., Baier W., Klein N., Strobel S. Oligosaccharides in human milk: structural, functional, and metabolic aspects. Annu Rev Nutr. 2000;20:699-722.

7. Hester S.N., Chen X., Li M., Monaco M.H., Comstock S.S., Kuhlenschmidt T.B., Kuhlenschmidt M.S., Donovan S.M. Human milk oligosaccharides inhibit rotavirus infectivity in vitro and in acutely infected piglets. Br J Nutr. 2013 Oct;110(7):1233-42.

8. Comstock S.S., Wang M., Hester S.N., Li M., Donovan S.M. Select human milk oligosaccharides directly modulate peripheral blood mononuclear cells isolated from 10-d-old pigs. Br J Nutr. 2014 Mar 14;111(5):819-28.

9. Comstock S.S., Li M., Wang M., Monaco M.H., Kuhlenschmidt T.B., Kuhlenschmidt M.S., Donovan S.M. Dietary human milk oligosaccharides but not prebiotic oligosaccharides increase circulating natural killer cell and mesenteric lymph node memory T cell populations in noninfected and rotavirus-infected neonatal piglets. J Nutr. 2017 Jun;147(6):1041-1047.

10. Li M., Monaco M.H., Wang M., Comstock S.S., Kuhlenschmidt T.B., Fahey G.C. Jr, Miller M.J., Kuhlenschmidt M.S., Donovan S.M. Human milk oligosaccharides shorten rotavirus-induced diarrhea and modulate piglet mucosal immunity and colonic microbiota. ISME J. 2014 Aug;8(8):1609-20.

 

Contributed by
Dr. Sandeep Ravindran
Freelance Science Writer
Sandeepr.com