Abstract
Objectives
Knowing that (i) high-fat diets upregulate inflammatory biomarkers in the small intestine and activate the mechanistic target of rapamycin complex 1 (mTORC1, a conserved protein kinase) and (ii) mTORC1 drives hepatic VLDL secretion, we investigated the role of intestinal mTORC1 in fat absorption and the lipid-regulating properties of 4-phenylbutyric acid (PBA) and R-α-lipoic acid (LA) in relation to mTORC1.
Methods
Lentivirus shRNA-based gene editing was utilized to create three stable Caco-2 cell lines that exhibit low, normal or high mTORC1 activity. Fat absorption was investigated in differentiated stable Caco-2 cells grown with oleate/albumin in Transwell permeable inserts.
Results
Genetic ablation of mTORC1 revealed that mTORC1 is required for the production and secretion of apoB-containing chylomicrons in Caco-2 cells. Rapamycin, an inhibitor of mTORC1, confirmed this finding as it lowered the secretion of apoB48 (−75%) and apoB100 (−47%) (P < 0.05). mTORC1 deletion was accompanied by the downregulation of APOB (−87%), MTTP (−94%), FASN (−76%), DGAT1 (−77%) and DGAT2 (−81%) genes (all P < 0.05); lower cellular abundance of apoB100 (−89%), MTTP (−63%) and FASN (−80%) (all P < 0.05); and lower media abundance of apoB48 (−66%), apoB100 (−51%) and triacylglycerols (−69%) (all P < 0.05). In Caco-2 cells harboring high mTORC1 activity, PBA or LA decreased the amounts of secreted apoB48 (−58% or −22%, respectively), apoB100 (−54% or −43%), and triacylglycerols (−46% or −38%), and the gene expression of APOB (−48% or −33%), MTTP (−54% or −31%), SREBF1 (−39% or −37%), FASN (−84% or −38%), and DGAT2 (−76% or −47%) (all P < 0.05).
Conclusions
A functional intestinal mTORC1 is indispensable for chylomicron production. Intake of short-chain fatty acid, PBA or LA, may be useful to prevent chylomicron overproduction-associated hypertriglyceridemia and postprandial inflammation.
Funding Sources
USDA Hatch Act, USDA-AFRI.
Electron microscopic study of intestinal fat absorption in vitro from mixed micelles containing linolenic acid, monoolein, and bile salt
