Milk fat derived FFA induces angiopoietin-like 4 gene
expression and protein secretion from intestinal cells
Jette F Young1, Søren D Nielsen1, Bashar Amer1, Grith Mortensen2, Bjørn Petrat-Melin1,
Randi Jessen1, Trine K Dalsgaard1
1
Department of Food Science, Aarhus University, 3380 Tjele, Denmark; 2Arla Foods, Strategic
Innovation Centre, 8220 Brabrand, Denmark
Key words: ANGPLT4 induction, free fatty acids, intestinal cell line, secretion
Background: In the 1980s, there was a warning against saturated fat and since then the public
perception of milk fat is to a certain extent negative because of its high content of saturated fatty
acids. However, an increasing number of studies have shown an inverse correlation between intake
of dairy products and both overweight and type 2 diabetes although this is not consistent. Recently,
by using an in vitro cell-based model we showed that whole milk up-regulated angiopoietin-like 4
(ANGPTL4) gene expression more than skimmed milk, indicating that milk fat may be responsible
for the ANGPTL4 gene induction. Free fatty acids (FFA) have previously been shown as major
regulators of ANGPLT4 gene expression and its derived protein is involved in the fat metabolism.
The ANGPTL4 protein has been suggested to suppress the clearance of circulating triglycerides and
to reduce the uptake of FFA within the surrounding tissue by inhibiting lipoprotein lipase (LPL)
activity, the enzyme hydrolyzing lipoprotein derived triglycerides in the blood stream.
Objective: To assess if FFA-derived from a milk-sample is capable of inducing the ANGPLT4
gene in an intestinal cell line and further investigate secretion of the protein in order to elucidate
possible mode of action.
Methods: The indigenous LPL in milk was used to generate elevated levels of FFA by
homogenising and incubating milk with serum which contains the LPL co-enzyme ApoCII. This
FFA-rich milk was added in a final concentration of 10% to an intestinal cell line (HCT 116) and
the ANGPLT4 gene expression was determined by real time qPCR. The intestinal cell line Caco-2
was seeded on permeable membranes and the fully differentiated cells were exposed to pure short
and medium chain FFA on the apical side. After incubation for 24 h the ANGPTL4 protein secretion
was determined from both the apical or basolateral side of the Caco-2 monolayer by sandwich ELISA.
Results: The level of FFA in whole milk correlated positively with ANGPTL4 gene expression in
the human colon cell line HCT 116. Highest induction of ANGPTL4 mRNA was observed in
response to medium chain FFA compared with short chain FFA, long chain saturated and
monounsaturated FFA. In a model system of differentiated Caco-2-cells seeded on permeable
membranes the FFA butyrate, caproate and caprylate induced ANGPTL4 gene expression and
further investigation of butyrate-induced ANGPTL4 regulation showed that protein was secreted to
both the apical and basolateral side of the Caco-2 monolayer, with the highest secretion to the apical
side.
Conclusion: Our study show that FFA of milk origin induce ANGPLT4 in a human intestinal cell
line and furthermore, that the protein is secreted to both the apical and basolateral side. This
indicate that ANGPLT4 may play a role in the uptake of fat from the small intestine as the
pancreatic lipase may be inhibited and hence limit FFA availability for intestinal uptake in a similar
way as inhibition of fat uptake from the blood stream to the different tissues.
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17th International Conference
Corresponding Author: Jette Feveile Young, PhD, Associate Professor, Institute of Food Science,
Aarhus University, 8830 Tjele, Denmark, E-mail: jettef.young@agrsci.dk , Phone number: (0045)
87158051, Secondary phone: (0045) 23478883
Main presenting author: Jette Feveile Young, PhD, Associate Professor, Institute of Food
Science, Aarhus University, 8830 Tjele, Denmark, E-mail: jettef.young@agrsci.dk , Phone number:
(0045) 87158051, Secondary phone: (0045) 23478883
Presentation type: Oral presentation
Main Conference Topics (please choose only one): Dairy functional foods
Co-authors:
Søren Drud Nielsen, PhD-student, E-mail: soren.drudnielsen@agrsci.dk
Bashar Amer, PhD, E-Mail: bashar.amer@agrsci.dk
Grith Mortensen, PhD, E-Mail: grith.mortensen@arlafoods.com
Bjørn Petrat-Melin, PhD-student, E-Mail: bjornm.nielsen@agrsci.dk
Randi Jessen, PhD, E-Mail: randi.jessen@agrsci.dk
Trine Kastrup Dalsgaard, PhD, E-mail: trine.dalsgaard@agrsci.dk