BACKGROUND & AIMS
Phosphatidylcholines (PCs) are structural and functional constituents of cell membranes. The activity of acyltransferase (LPCAT) is required for addition of polyunsaturated fatty acids to the sn-2 position of PCs, and is therefore required to maintain cell membrane structure and function. LPCAT3 is the most abundant isoform of LPCAT in the small intestine and liver—are important sites of plasma lipoprotein metabolism. We investigated the effects of Lpcat3 disruption on lipid metabolism in mice.
METHODS
We disrupted the gene Lpcat3 in C57BL/6J mice to create LPCAT3 knockout (KO) mice. Livers and small intestinal tissues were collected from LPCAT3 KO and C57BL/6J parental strain (controls), and levels of LPCAT mRNAs and protein were measured. Levels of lipids and lipoproteins were measured in plasma samples. We isolated enterocytes from mice and measured levels of RNAs and proteins involved in lipid uptake by real-time PCR and immunoblot assays, respectively. We assessed lipid absorption and PC subspecies in the enterocyte plasma membrane using liquid chromatography with tandem mass spectometry.
RESULTS
LPCAT3 KO mice survived only 3 weeks after birth. Oil Red O staining showed that the control but not LPCAT3 KO mice accumulated lipids in the small intestine; levels of Niemann-Pick C1-like 1 (NPC1L1) and fatty acid transporter protein 4 (FATP4), which regulate lipid uptake, were greatly reduced in the small intestines of LPCAT3 KO mice. Oral administration of PC and olive oil allowed the LPCAT3 KO mice to survive, with the same body weights as controls, but the KO mice had shorter and wider small-intestinal villi and longer and bigger small intestines. Plasma membranes of enterocytes from LPCAT3 KO mice also had significant reductions in the composition of polyunsaturated PCs and reduced levels of NPC1L1, CD36, and FATP4 proteins. These reductions were associated with reduced intestinal uptake of lipid by the small intestine and reduced plasma levels of cholesterol, phospholipid, and triglyceride.
CONCLUSIONS
LPCAT3 KO mice have longer and larger small intestines than control mice, with shorter wide villi, reduced lipid absorption, and lower levels NPC1L1, CD36, and FATP4 proteins. Inhibition of LPCAT3 in the small intestine could be developed as an approach to treat hyperlipidemia.
Serine palmitoyltransferase (SPT) is the first and rate-limiting enzyme of the de novo biosynthetic pathway of sphingomyelin (SM). Both SPT and SM have been implicated in the pathogenesis of atherosclerosis, the development of which is driven by macrophages; however, the role of SPT in macrophage-mediated atherogenesis is unknown. To address this issue, we have analyzed macrophage inflammatory responses and reverse cholesterol transport, 2 key mediators of atherogenesis, in SPT subunit 2-haploinsufficient (Sptlc2 +/-) macrophages. We found that Sptlc2 +/-macrophages have significantly lower SM levels in plasma membrane and lipid rafts. This reduction not only impaired inflammatory responses triggered by TLR4 and its downstream NF-κB and MAPK pathways, but also enhanced reverse cholesterol transport mediated by ABC transporters. LDL receptor-deficient (Ldlr -/-) mice transplanted with Sptlc2 +/-bone marrow cells exhibited significantly fewer atherosclerotic lesions after high-fat and high-cholesterol diet feeding. Additionally, Ldlr -/-mice with myeloid cell-specific Sptlc2 haploinsufficiency exhibited significantly less atherosclerosis than controls. These findings suggest that SPT could be a novel therapeutic target in atherosclerosis.
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