Chenyuan Wang1,2, Ming Zhang3, Huiyuan Guo1,2, Jingyu Yan4, Lingli Chen1,2, Wendi Teng1,2, Fazheng Ren1,2, Yiran Li1,2, Xifan Wang1,2, Jie Luo5, Yixuan Li1. 1. Beijing Advanced Innovation Center for Food Nutrition and Human Health, College of Food Science & Nutritional Engineering, China Agricultural University, Beijing, China. 2. Key Laboratory of Functional Dairy, China Agricultural University, Beijing, China. 3. School of Food and Chemical Engineering, Beijing Technology and Business University, Beijing, China. 4. Dalian Institute of Chemical Physics, Chinese Academy of Sciences, Dalian, China. 5. College of Food Science and Technology, Hunan Agricultural University, Changsha, China.
Abstract
BACKGROUND: Hypoxia-induced intestinal barrier injuries lead to necrotizing enterocolitis (NEC). Although NEC in preterm neonates is preventable by human milk oligosaccharides (HMOs), the underlying mechanism remains unknown. OBJECTIVE: To reveal the role and mechanism of HMOs in protecting against hypoxia-induced injuries in intestinal epithelium of neonatal mice and cultured Caco2 cells. METHODS: NEC was induced by hypoxia and cold stress. Seventy C57BL/C pups (7-d-old) were divided into 5 groups and fed maternal breast milk (BM), formula alone (FF), or the formula added with HMOs at 5 (LHMO), 10 (MHMO), or 20 mg/mL (HHMO) for 3 d. Ileal hypoxia inducible factor 1α (HIF1α) and cleaved Caspase 3 were determined, along with staining for Ki-67 protein to labeled proliferative cells. In vitro, adherent Caco2 cells (undifferentiated, passage 14) were treated with HMOs, galacto-oligosaccharides, fructo-oligosaccharides, or mixed oligosaccharides at 10 mg/mL for 1 d exposed to 1% O2. Cell proliferation and apoptosis, along with phosphorylated epidermal growth factor receptor (P-EGFR) and 38KD MAPK (P-P38), were assayed in differentiated or undifferentiated Caco2 cells. RESULTS: Compared with the FF-fed mice, those fed MHMO and HHMO had 52% lower (P < 0.05) NEC scores, 60-80% greater (P < 0.05) KI67-positive cell numbers, and 56-71% decreases (P < 0.05) in ileal HIF1α and cleaved Caspase 3 (56-71%). Compared with those untreated, the HMO-treated Caco2 cells displayed 60% greater (P < 0.05) proliferative activity and 19% lower (P < 0.05) apoptotic cells after the hypoxia exposure. The HMO treatment led to 58% or 10-fold increases (P < 0.05) of P-EGFR and 48-89% decreases (P < 0.05) of P-P38 in either differentiated or undifferentiated Caco2 cells compared with the controls. CONCLUSION: Supplementing HMOs at 10-20 mg/mL into the formula for neonatal mice or media for Caco2 cells conferred protection against the hypoxia-induced injuries. The protection in the Caco2 cells was associated with an activation of EGFR.
BACKGROUND:Hypoxia-induced intestinal barrier injuries lead to necrotizing enterocolitis (NEC). Although NEC in preterm neonates is preventable by human milk oligosaccharides (HMOs), the underlying mechanism remains unknown. OBJECTIVE: To reveal the role and mechanism of HMOs in protecting against hypoxia-induced injuries in intestinal epithelium of neonatal mice and cultured Caco2 cells. METHODS:NEC was induced by hypoxia and cold stress. Seventy C57BL/C pups (7-d-old) were divided into 5 groups and fed maternal breast milk (BM), formula alone (FF), or the formula added with HMOs at 5 (LHMO), 10 (MHMO), or 20 mg/mL (HHMO) for 3 d. Ileal hypoxia inducible factor 1α (HIF1α) and cleaved Caspase 3 were determined, along with staining for Ki-67 protein to labeled proliferative cells. In vitro, adherent Caco2 cells (undifferentiated, passage 14) were treated with HMOs, galacto-oligosaccharides, fructo-oligosaccharides, or mixed oligosaccharides at 10 mg/mL for 1 d exposed to 1% O2. Cell proliferation and apoptosis, along with phosphorylated epidermal growth factor receptor (P-EGFR) and 38KD MAPK (P-P38), were assayed in differentiated or undifferentiated Caco2 cells. RESULTS: Compared with the FF-fed mice, those fed MHMO and HHMO had 52% lower (P < 0.05) NEC scores, 60-80% greater (P < 0.05) KI67-positive cell numbers, and 56-71% decreases (P < 0.05) in ileal HIF1α and cleaved Caspase 3 (56-71%). Compared with those untreated, the HMO-treated Caco2 cells displayed 60% greater (P < 0.05) proliferative activity and 19% lower (P < 0.05) apoptotic cells after the hypoxia exposure. The HMO treatment led to 58% or 10-fold increases (P < 0.05) of P-EGFR and 48-89% decreases (P < 0.05) of P-P38 in either differentiated or undifferentiated Caco2 cells compared with the controls. CONCLUSION: Supplementing HMOs at 10-20 mg/mL into the formula for neonatal mice or media for Caco2 cells conferred protection against the hypoxia-induced injuries. The protection in the Caco2 cells was associated with an activation of EGFR.
Authors: Ilse H de Lange; Charlotte van Gorp; Laurens D Eeftinck Schattenkerk; Wim G van Gemert; Joep P M Derikx; Tim G A M Wolfs Journal: Nutrients Date: 2021-05-19 Impact factor: 5.717