PMID- 32221473 OWN - NLM STAT- MEDLINE DCOM- 20211122 LR - 20211122 IS - 1530-0447 (Electronic) IS - 0031-3998 (Print) IS - 0031-3998 (Linking) VI - 89 IP - 1 DP - 2021 Jan TI - The human milk oligosaccharides 2'-fucosyllactose and 6'-sialyllactose protect against the development of necrotizing enterocolitis by inhibiting toll-like receptor 4 signaling. PG - 91-101 LID - 10.1038/s41390-020-0852-3 [doi] AB - BACKGROUND: Necrotizing enterocolitis (NEC) develops through exaggerated toll-like receptor 4 (TLR4) signaling in the intestinal epithelium. Breast milk is rich in non-digestible oligosaccharides and prevents NEC through unclear mechanisms. We now hypothesize that the human milk oligosaccharides 2'-fucosyllactose (2'-FL) and 6'-sialyllactose (6'-SL) can reduce NEC through inhibition of TLR4 signaling. METHODS: NEC was induced in newborn mice and premature piglets and infant formula was supplemented with 2'-FL, 6'-SL, or lactose. Intestinal tissue was obtained at surgical resection. HMO inhibition of TLR4 was assessed in IEC-6 enterocytes, mice, and human tissue explants and via in silico modeling. RESULTS: Supplementation of infant formula with either 2'-FL and/or 6'-SL, but not the parent sugar lactose, reduced NEC in mice and piglets via reduced apoptosis, inflammation, weight loss, and histological appearance. Mechanistically, both 2'-FL and 6'-SL, but not lactose, reduced TLR4-mediated nuclear factor kappa light-chain enhancer of activated B cells (NF-kB) inflammatory signaling in the mouse and human intestine. Strikingly, in silico modeling revealed 2'-FL and 6'-SL, but not lactose, to dock into the binding pocket of the TLR4-MD2 complex, explaining their ability to inhibit TLR4 signaling. CONCLUSIONS: 2'-FL and 6'-SL, but not lactose, prevent NEC in mice and piglet models and attenuate NEC inflammation in the human ileum, in part through TLR4 inhibition. IMPACT: Necrotizing enterocolitis (NEC) is a major cause of morbidity and mortality in premature infants that occurs in the setting of bacterial colonization of the gut and administration of formula feeds and activation by the innate immune receptor toll-like receptor 4 (TLR4). Breast milk prevents NEC through unclear mechanisms. We now show that breast milk-enriched human milk oligosaccharides (HMOs) that are derived from lactose prevent NEC through inhibition of TLR4. The human milk oligosaccharides 2'-FL and 6'-SL, but not the backbone sugar lactose, prevent NEC in mice and piglets. 2'-FL and 6'-SL but not lactose inhibited TLR4 signaling in cultured enterocytes, in enteroids derived from mouse intestine, and in human intestinal explants obtained at the time of surgical resection for patients with NEC. In seeking the mechanisms involved, 2'-FL and 6'-SL but not lactose were found to directly bind to TLR4, explaining the inhibition and protection against NEC. These findings may impact clinical practice by suggesting that administration of HMOs could serve as a preventive strategy for premature infants at risk for NEC development. FAU - Sodhi, Chhinder P AU - Sodhi CP AD - Division of General Pediatric Surgery, Johns Hopkins University and Johns Hopkins Children's Center, Baltimore, MD, 21287, USA. AD - Department of Surgery, Johns Hopkins University and Johns Hopkins Children's Center, Baltimore, MD, 21287, USA. FAU - Wipf, Peter AU - Wipf P AD - Department of Chemistry, University of Pittsburgh, Pittsburgh, PA, USA. FAU - Yamaguchi, Yukihiro AU - Yamaguchi Y AD - Division of General Pediatric Surgery, Johns Hopkins University and Johns Hopkins Children's Center, Baltimore, MD, 21287, USA. AD - Department of Surgery, Johns Hopkins University and Johns Hopkins Children's Center, Baltimore, MD, 21287, USA. FAU - Fulton, William B AU - Fulton WB AD - Division of General Pediatric Surgery, Johns Hopkins University and Johns Hopkins Children's Center, Baltimore, MD, 21287, USA. AD - Department of Surgery, Johns Hopkins University and Johns Hopkins Children's Center, Baltimore, MD, 21287, USA. FAU - Kovler, Mark AU - Kovler M AD - Division of General Pediatric Surgery, Johns Hopkins University and Johns Hopkins Children's Center, Baltimore, MD, 21287, USA. AD - Department of Surgery, Johns Hopkins University and Johns Hopkins Children's Center, Baltimore, MD, 21287, USA. FAU - Nino, Diego F AU - Nino DF AD - Division of General Pediatric Surgery, Johns Hopkins University and Johns Hopkins Children's Center, Baltimore, MD, 21287, USA. AD - Department of Surgery, Johns Hopkins University and Johns Hopkins Children's Center, Baltimore, MD, 21287, USA. FAU - Zhou, Qinjie AU - Zhou Q AD - Division of General Pediatric Surgery, Johns Hopkins University and Johns Hopkins Children's Center, Baltimore, MD, 21287, USA. AD - Department of Surgery, Johns Hopkins University and Johns Hopkins Children's Center, Baltimore, MD, 21287, USA. FAU - Banfield, Emilyn AU - Banfield E AD - Division of General Pediatric Surgery, Johns Hopkins University and Johns Hopkins Children's Center, Baltimore, MD, 21287, USA. FAU - Werts, Adam D AU - Werts AD AD - Department of Chemistry, University of Pittsburgh, Pittsburgh, PA, USA. FAU - Ladd, Mitchell R AU - Ladd MR AD - Division of General Pediatric Surgery, Johns Hopkins University and Johns Hopkins Children's Center, Baltimore, MD, 21287, USA. AD - Department of Surgery, Johns Hopkins University and Johns Hopkins Children's Center, Baltimore, MD, 21287, USA. FAU - Buck, Rachael H AU - Buck RH AD - Abbott Nutrition, a Division of Abbott Laboratories, Columbus, OH, USA. FAU - Goehring, Karen C AU - Goehring KC AD - Abbott Nutrition, a Division of Abbott Laboratories, Columbus, OH, USA. FAU - Prindle, Thomas Jr AU - Prindle T Jr AD - Division of General Pediatric Surgery, Johns Hopkins University and Johns Hopkins Children's Center, Baltimore, MD, 21287, USA. AD - Department of Surgery, Johns Hopkins University and Johns Hopkins Children's Center, Baltimore, MD, 21287, USA. FAU - Wang, Sanxia AU - Wang S AD - Division of General Pediatric Surgery, Johns Hopkins University and Johns Hopkins Children's Center, Baltimore, MD, 21287, USA. AD - Department of Surgery, Johns Hopkins University and Johns Hopkins Children's Center, Baltimore, MD, 21287, USA. FAU - Jia, Hongpeng AU - Jia H AD - Division of General Pediatric Surgery, Johns Hopkins University and Johns Hopkins Children's Center, Baltimore, MD, 21287, USA. AD - Department of Surgery, Johns Hopkins University and Johns Hopkins Children's Center, Baltimore, MD, 21287, USA. FAU - Lu, Peng AU - Lu P AD - Division of General Pediatric Surgery, Johns Hopkins University and Johns Hopkins Children's Center, Baltimore, MD, 21287, USA. AD - Department of Surgery, Johns Hopkins University and Johns Hopkins Children's Center, Baltimore, MD, 21287, USA. FAU - Hackam, David J AU - Hackam DJ AUID- ORCID: 0000-0001-5195-8807 AD - Division of General Pediatric Surgery, Johns Hopkins University and Johns Hopkins Children's Center, Baltimore, MD, 21287, USA. dhackam1@jhmi.edu. AD - Department of Surgery, Johns Hopkins University and Johns Hopkins Children's Center, Baltimore, MD, 21287, USA. dhackam1@jhmi.edu. LA - eng GR - R01 DK117186/DK/NIDDK NIH HHS/United States GR - R01 DK083752/DK/NIDDK NIH HHS/United States GR - R01 GM078238/GM/NIGMS NIH HHS/United States GR - T32 DK007713/DK/NIDDK NIH HHS/United States GR - T32 OD011089/OD/NIH HHS/United States PT - Journal Article PT - Research Support, N.I.H., Extramural PT - Research Support, Non-U.S. Gov't DEP - 20200327 PL - United States TA - Pediatr Res JT - Pediatric research JID - 0100714 RN - 0 (6'-sialyllactose) RN - 0 (Inflammation Mediators) RN - 0 (TLR4 protein, human) RN - 0 (Tlr4 protein, mouse) RN - 0 (Toll-Like Receptor 4) RN - 0 (Trisaccharides) RN - J2B2A4N98G (Lactose) RN - XO2533XO8R (2'-fucosyllactose) SB - IM CIN - Pediatr Res. 2021 Jan;89(1):248. PMID: 32998155 MH - Animals MH - Animals, Newborn MH - Apoptosis/drug effects MH - Cell Line MH - Disease Models, Animal MH - Enterocolitis, Necrotizing/immunology/metabolism/pathology/*prevention & control MH - Humans MH - Ileum/*drug effects/immunology/metabolism/pathology MH - Inflammation Mediators/metabolism MH - Intestinal Mucosa/*drug effects/immunology/metabolism/pathology MH - Lactose/*analogs & derivatives/isolation & purification/pharmacology MH - Mice MH - Milk, Human/*chemistry MH - Molecular Docking Simulation MH - Signal Transduction MH - Sus scrofa MH - Toll-Like Receptor 4/*antagonists & inhibitors/metabolism MH - Trisaccharides/isolation & purification/*pharmacology MH - Weight Loss/drug effects PMC - PMC7529714 MID - NIHMS1575938 EDAT- 2020/03/30 06:00 MHDA- 2021/11/23 06:00 PMCR- 2021/02/14 CRDT- 2020/03/30 06:00 PHST- 2019/08/28 00:00 [received] PHST- 2020/02/19 00:00 [accepted] PHST- 2020/02/10 00:00 [revised] PHST- 2020/03/30 06:00 [pubmed] PHST- 2021/11/23 06:00 [medline] PHST- 2020/03/30 06:00 [entrez] PHST- 2021/02/14 00:00 [pmc-release] AID - 10.1038/s41390-020-0852-3 [pii] AID - 10.1038/s41390-020-0852-3 [doi] PST - ppublish SO - Pediatr Res. 2021 Jan;89(1):91-101. doi: 10.1038/s41390-020-0852-3. Epub 2020 Mar 27.