title: Microlipid-induced oxidative stress in human breastmilk: in vitro effects on intestinal epithelial cells.
abstract: OBJECTIVE
  To (1) determine whether medium chain fatty acids (Microlipid) added to human breastmilk generates reactive oxygen species (ROS), and (2) measure the physiological effect(s) of Microlipid) (ML)-supplemented human breastmilk in an enterocyte cell culture bioassay.
  
  
  METHODS
  ML was added to milk according to manufacturer's recommendations and total hydroperoxides measured at intervals with the FOX 2 and TBARS assays. Physiological effects of supplementation were measured using a human enterocyte cell line (Caco-2BBE) and/or a primary human fetal intestinal cell culture (FHS-74 Int). Endpoints included: intracellular oxidative stress, transepithelial electrical resistance (TEER), apoptosis, and interleukin (IL)-6 production.
  
  
  RESULTS
  Immediately postsupplementation, ML did not significantly increase ROS, as determined by both the FOX 2 and TBARS assays. Further, storage of milk + ML at 4 degrees C prevented significant increases in total hydroperoxides. However, by 4 hours postsupplementation at room temperature, both assays revealed significantly higher hydroperoxide and lipid peroxide levels. ML-supplemented milk stored at room temperature for 4 hours had the following effects in cell culture bioassays: elevated oxidative stress, increased rates of apoptosis, decreased transmembrane electrical resistance (TEER) values and, in both cell culture assays, significantly increased secretion of IL-6.
  
  
  CONCLUSIONS
  Based on our measurements of extracellular and intracellular ROS, milk supplemented with fresh ML does not induce significant oxidative stress. However, when stored for 4 hours at room temperature, ML induces significant levels of oxidative stress. Decreases in TEER and increases in apoptosis and IL-6 secretion are consistent with ML-induced oxidative stress. It therefore is likely that in clinical situations, if ML-supplemented milk is not administered quickly, the newborn may be placed at greater risk of oxidative stress.
authors: Faculty of Nursing, University of Manitoba, Winnipeg, Manitoba, Canada. Bill_Diehl-Jones@umanitoba.ca
authors: William L
authors: WL
authors: Diehl-Jones
authors: Debra Fraser
authors: DF
authors: Askin
authors: James K
authors: JK
authors: Friel
mesh_terms: D000704:Analysis of Variance; D017209:Apoptosis / Q000187:drug effects; D001681:Biological Assay; D018938:Caco-2 Cells; D020895:Enterocytes / Q000378:metabolism*; D005227:Fatty Acids / Q000494:pharmacology*; D005519:Food Preservation / Q000379:methods; D006801:Humans; D006861:Hydrogen Peroxide / Q000032:analysis; D015850:Interleukin-6 / Q000378:metabolism; D015227:Lipid Peroxidation / Q000187:drug effects; D008895:Milk, Human / Q000276:immunology*; D018384:Oxidative Stress / Q000187:drug effects*; D017382:Reactive Oxygen Species / Q000032:analysis*; D013696:Temperature; D017392:Thiobarbituric Acid Reactive Substances / Q000032:analysis; D013997:Time Factors
chemical_list: D005227:Fatty Acids; D015850:Interleukin-6; D017382:Reactive Oxygen Species; D017392:Thiobarbituric Acid Reactive Substances; D006861:Hydrogen Peroxide