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Lpcat3-dependent production of arachidonoyl phospholipids is a key determinant of triglyceride secretion.

Rong X, Wang B, Dunham MM, Hedde PN, Wong JS, Gratton E, Young SG, Ford DA, Tontonoz P - Elife (2015)

Bottom Line: Mice lacking Lpcat3 in the intestine fail to thrive during weaning and exhibit enterocyte lipid accumulation and reduced plasma TGs.Mice lacking Lpcat3 in the liver show reduced plasma TGs, hepatosteatosis, and secrete lipid-poor very low-density lipoprotein (VLDL) lacking arachidonoyl PLs.Mechanistic studies indicate that Lpcat3 activity impacts membrane lipid mobility in living cells, suggesting a biophysical basis for the requirement of arachidonoyl PLs in lipidating lipoprotein particles.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology and Laboratory Medicine, Howard Hughes Medical Institute, University of California, Los Angeles, Los Angeles, United States.

ABSTRACT
The role of specific phospholipids (PLs) in lipid transport has been difficult to assess due to an inability to selectively manipulate membrane composition in vivo. Here we show that the phospholipid remodeling enzyme lysophosphatidylcholine acyltransferase 3 (Lpcat3) is a critical determinant of triglyceride (TG) secretion due to its unique ability to catalyze the incorporation of arachidonate into membranes. Mice lacking Lpcat3 in the intestine fail to thrive during weaning and exhibit enterocyte lipid accumulation and reduced plasma TGs. Mice lacking Lpcat3 in the liver show reduced plasma TGs, hepatosteatosis, and secrete lipid-poor very low-density lipoprotein (VLDL) lacking arachidonoyl PLs. Mechanistic studies indicate that Lpcat3 activity impacts membrane lipid mobility in living cells, suggesting a biophysical basis for the requirement of arachidonoyl PLs in lipidating lipoprotein particles. These data identify Lpcat3 as a key factor in lipoprotein production and illustrate how manipulation of membrane composition can be used as a regulatory mechanism to control metabolic pathways.

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Altered TG metabolism in intestine-specific Lpcat3 knockout mice.(A) Induction of Lpcat3 mRNA expression in duodenum of mice treated with 40 mg/kg/day GW3956 by oral gavage for 3 days (n = 5/group). Gene expression was measured by real-time PCR. Values are means ± SEM. (B) Representative photograph and body weight of newborn Lpcat3fl/flVillin-cre (IKO) and control Lpcat3fl/fl (F/F) pups (n = 5/group for body weight measurement). Values are means ± SEM. (C) Representative photograph and body weight of 1 week-old Lpcat3fl/flVillin-cre (IKO) and control Lpcat3fl/fl (F/F) pups (n ≥ 6/group for body weight measurement). Values are means ± SEM. (D) Expression of Lpcat family members in 1 week-old Lpcat3fl/fl (F/F) and Lpcat3fl/flVillin-cre (IKO) duodenum measured by real-time PCR (n ≥ 6/group). Ct values of F/F samples were shown. Values are means ± SEM. (E) Blood glucose, plasma lipids and insulin levels in 1 week-old Lpcat3fl/fl (F/F) and Lpcat3fl/flVillin-cre (IKO) pups (n ≥ 6/group). Values are means ± SEM. (F) Hematoxylin and eosin staining of intestines from 1 week-old Lpcat3fl/fl (WT) and Lpcat3fl/flVillin-cre (IKO) pups. (G) Expression of genes in duodenum of 1 week-old Lpcat3fl/fl (WT) and Lpcat3fl/flVillin-cre (IKO) pups. Gene expression was measured by real-time PCR (n ≥ 6/group). Values are means ± SEM. Statistical analysis was performed using Student's t-test (A, B, D, E and F). *p < 0.05; **p < 0.01.DOI:http://dx.doi.org/10.7554/eLife.06557.011
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fig4: Altered TG metabolism in intestine-specific Lpcat3 knockout mice.(A) Induction of Lpcat3 mRNA expression in duodenum of mice treated with 40 mg/kg/day GW3956 by oral gavage for 3 days (n = 5/group). Gene expression was measured by real-time PCR. Values are means ± SEM. (B) Representative photograph and body weight of newborn Lpcat3fl/flVillin-cre (IKO) and control Lpcat3fl/fl (F/F) pups (n = 5/group for body weight measurement). Values are means ± SEM. (C) Representative photograph and body weight of 1 week-old Lpcat3fl/flVillin-cre (IKO) and control Lpcat3fl/fl (F/F) pups (n ≥ 6/group for body weight measurement). Values are means ± SEM. (D) Expression of Lpcat family members in 1 week-old Lpcat3fl/fl (F/F) and Lpcat3fl/flVillin-cre (IKO) duodenum measured by real-time PCR (n ≥ 6/group). Ct values of F/F samples were shown. Values are means ± SEM. (E) Blood glucose, plasma lipids and insulin levels in 1 week-old Lpcat3fl/fl (F/F) and Lpcat3fl/flVillin-cre (IKO) pups (n ≥ 6/group). Values are means ± SEM. (F) Hematoxylin and eosin staining of intestines from 1 week-old Lpcat3fl/fl (WT) and Lpcat3fl/flVillin-cre (IKO) pups. (G) Expression of genes in duodenum of 1 week-old Lpcat3fl/fl (WT) and Lpcat3fl/flVillin-cre (IKO) pups. Gene expression was measured by real-time PCR (n ≥ 6/group). Values are means ± SEM. Statistical analysis was performed using Student's t-test (A, B, D, E and F). *p < 0.05; **p < 0.01.DOI:http://dx.doi.org/10.7554/eLife.06557.011

Mentions: Lpcat3 is expressed at high levels in intestine as well as in the liver. We showed previously that hepatic Lpcat3 expression is regulated by the sterol-activated nuclear receptor LXR (Rong et al., 2013). Here, we showed that intestinal Lpcat3 expression is strongly responsive to the administration of a synthetic LXR-agonist, GW3965 (Figure 4A). To address whether Lpcat3 activity may also be important for TG metabolism in intestinal enterocytes, we generated intestine-specific Lpcat3 KO mice (I-Lpcat3 KO) by crossing the floxed mice to villin-Cre transgenics. I-Lpcat3 KO mice were born at the predicted Mendelian frequency, and their body weights at birth were comparable to controls (Table 3, Figure 4B). However, even though the pups suckled, they failed to thrive and showed severe growth retardation by 1 week of age (Figure 4C). Expression of Lpcat3 was reduced more than 90% in duodenum of I-Lpcat3 KO mice as expected, and there was no compensatory increase in expression of Lpcat1, Lpcat2 or Lpcat4 (Figure 4D). Blood glucose levels in 1-week-old I-Lpcat3 pups were very low (Figure 4E), consistent with results obtained with global knockouts (Figure 1). Plasma insulin levels were also correspondingly reduced. Plasma TG levels were lower and total cholesterol and NEFA levels were unchanged in I-Lpcat3 KO pups (Figure 4E). Histological analysis of intestines from I-Lpcat3 KO pups revealed a dramatic accumulation of cytosolic lipid droplets in intestinal enterocytes (Figure 4F), a phenotype reminiscent of intestinal apoB-deficient mice. Analysis of intestinal gene expression in I-Lpcat3 KO mice revealed reduced expression of several genes linked to intestinal TG metabolism, including Apob, Cd36, Dgat2, and Mogat2 (Figure 4G). Given the massive enterocyte lipid accumulation in enterocytes, it is conceivable that some of those gene-expression changes were due, at least in part, to poor nutrition or cell toxicity. Nevertheless, these data were consistent with a role for Lpcat3 in TG mobilization and secretion–in the intestine as well as in the liver.10.7554/eLife.06557.011Figure 4.Altered TG metabolism in intestine-specific Lpcat3 knockout mice.


Lpcat3-dependent production of arachidonoyl phospholipids is a key determinant of triglyceride secretion.

Rong X, Wang B, Dunham MM, Hedde PN, Wong JS, Gratton E, Young SG, Ford DA, Tontonoz P - Elife (2015)

Altered TG metabolism in intestine-specific Lpcat3 knockout mice.(A) Induction of Lpcat3 mRNA expression in duodenum of mice treated with 40 mg/kg/day GW3956 by oral gavage for 3 days (n = 5/group). Gene expression was measured by real-time PCR. Values are means ± SEM. (B) Representative photograph and body weight of newborn Lpcat3fl/flVillin-cre (IKO) and control Lpcat3fl/fl (F/F) pups (n = 5/group for body weight measurement). Values are means ± SEM. (C) Representative photograph and body weight of 1 week-old Lpcat3fl/flVillin-cre (IKO) and control Lpcat3fl/fl (F/F) pups (n ≥ 6/group for body weight measurement). Values are means ± SEM. (D) Expression of Lpcat family members in 1 week-old Lpcat3fl/fl (F/F) and Lpcat3fl/flVillin-cre (IKO) duodenum measured by real-time PCR (n ≥ 6/group). Ct values of F/F samples were shown. Values are means ± SEM. (E) Blood glucose, plasma lipids and insulin levels in 1 week-old Lpcat3fl/fl (F/F) and Lpcat3fl/flVillin-cre (IKO) pups (n ≥ 6/group). Values are means ± SEM. (F) Hematoxylin and eosin staining of intestines from 1 week-old Lpcat3fl/fl (WT) and Lpcat3fl/flVillin-cre (IKO) pups. (G) Expression of genes in duodenum of 1 week-old Lpcat3fl/fl (WT) and Lpcat3fl/flVillin-cre (IKO) pups. Gene expression was measured by real-time PCR (n ≥ 6/group). Values are means ± SEM. Statistical analysis was performed using Student's t-test (A, B, D, E and F). *p < 0.05; **p < 0.01.DOI:http://dx.doi.org/10.7554/eLife.06557.011
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fig4: Altered TG metabolism in intestine-specific Lpcat3 knockout mice.(A) Induction of Lpcat3 mRNA expression in duodenum of mice treated with 40 mg/kg/day GW3956 by oral gavage for 3 days (n = 5/group). Gene expression was measured by real-time PCR. Values are means ± SEM. (B) Representative photograph and body weight of newborn Lpcat3fl/flVillin-cre (IKO) and control Lpcat3fl/fl (F/F) pups (n = 5/group for body weight measurement). Values are means ± SEM. (C) Representative photograph and body weight of 1 week-old Lpcat3fl/flVillin-cre (IKO) and control Lpcat3fl/fl (F/F) pups (n ≥ 6/group for body weight measurement). Values are means ± SEM. (D) Expression of Lpcat family members in 1 week-old Lpcat3fl/fl (F/F) and Lpcat3fl/flVillin-cre (IKO) duodenum measured by real-time PCR (n ≥ 6/group). Ct values of F/F samples were shown. Values are means ± SEM. (E) Blood glucose, plasma lipids and insulin levels in 1 week-old Lpcat3fl/fl (F/F) and Lpcat3fl/flVillin-cre (IKO) pups (n ≥ 6/group). Values are means ± SEM. (F) Hematoxylin and eosin staining of intestines from 1 week-old Lpcat3fl/fl (WT) and Lpcat3fl/flVillin-cre (IKO) pups. (G) Expression of genes in duodenum of 1 week-old Lpcat3fl/fl (WT) and Lpcat3fl/flVillin-cre (IKO) pups. Gene expression was measured by real-time PCR (n ≥ 6/group). Values are means ± SEM. Statistical analysis was performed using Student's t-test (A, B, D, E and F). *p < 0.05; **p < 0.01.DOI:http://dx.doi.org/10.7554/eLife.06557.011
Mentions: Lpcat3 is expressed at high levels in intestine as well as in the liver. We showed previously that hepatic Lpcat3 expression is regulated by the sterol-activated nuclear receptor LXR (Rong et al., 2013). Here, we showed that intestinal Lpcat3 expression is strongly responsive to the administration of a synthetic LXR-agonist, GW3965 (Figure 4A). To address whether Lpcat3 activity may also be important for TG metabolism in intestinal enterocytes, we generated intestine-specific Lpcat3 KO mice (I-Lpcat3 KO) by crossing the floxed mice to villin-Cre transgenics. I-Lpcat3 KO mice were born at the predicted Mendelian frequency, and their body weights at birth were comparable to controls (Table 3, Figure 4B). However, even though the pups suckled, they failed to thrive and showed severe growth retardation by 1 week of age (Figure 4C). Expression of Lpcat3 was reduced more than 90% in duodenum of I-Lpcat3 KO mice as expected, and there was no compensatory increase in expression of Lpcat1, Lpcat2 or Lpcat4 (Figure 4D). Blood glucose levels in 1-week-old I-Lpcat3 pups were very low (Figure 4E), consistent with results obtained with global knockouts (Figure 1). Plasma insulin levels were also correspondingly reduced. Plasma TG levels were lower and total cholesterol and NEFA levels were unchanged in I-Lpcat3 KO pups (Figure 4E). Histological analysis of intestines from I-Lpcat3 KO pups revealed a dramatic accumulation of cytosolic lipid droplets in intestinal enterocytes (Figure 4F), a phenotype reminiscent of intestinal apoB-deficient mice. Analysis of intestinal gene expression in I-Lpcat3 KO mice revealed reduced expression of several genes linked to intestinal TG metabolism, including Apob, Cd36, Dgat2, and Mogat2 (Figure 4G). Given the massive enterocyte lipid accumulation in enterocytes, it is conceivable that some of those gene-expression changes were due, at least in part, to poor nutrition or cell toxicity. Nevertheless, these data were consistent with a role for Lpcat3 in TG mobilization and secretion–in the intestine as well as in the liver.10.7554/eLife.06557.011Figure 4.Altered TG metabolism in intestine-specific Lpcat3 knockout mice.

Bottom Line: Mice lacking Lpcat3 in the intestine fail to thrive during weaning and exhibit enterocyte lipid accumulation and reduced plasma TGs.Mice lacking Lpcat3 in the liver show reduced plasma TGs, hepatosteatosis, and secrete lipid-poor very low-density lipoprotein (VLDL) lacking arachidonoyl PLs.Mechanistic studies indicate that Lpcat3 activity impacts membrane lipid mobility in living cells, suggesting a biophysical basis for the requirement of arachidonoyl PLs in lipidating lipoprotein particles.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology and Laboratory Medicine, Howard Hughes Medical Institute, University of California, Los Angeles, Los Angeles, United States.

ABSTRACT
The role of specific phospholipids (PLs) in lipid transport has been difficult to assess due to an inability to selectively manipulate membrane composition in vivo. Here we show that the phospholipid remodeling enzyme lysophosphatidylcholine acyltransferase 3 (Lpcat3) is a critical determinant of triglyceride (TG) secretion due to its unique ability to catalyze the incorporation of arachidonate into membranes. Mice lacking Lpcat3 in the intestine fail to thrive during weaning and exhibit enterocyte lipid accumulation and reduced plasma TGs. Mice lacking Lpcat3 in the liver show reduced plasma TGs, hepatosteatosis, and secrete lipid-poor very low-density lipoprotein (VLDL) lacking arachidonoyl PLs. Mechanistic studies indicate that Lpcat3 activity impacts membrane lipid mobility in living cells, suggesting a biophysical basis for the requirement of arachidonoyl PLs in lipidating lipoprotein particles. These data identify Lpcat3 as a key factor in lipoprotein production and illustrate how manipulation of membrane composition can be used as a regulatory mechanism to control metabolic pathways.

Show MeSH
Related in: MedlinePlus