Chronic overexpression of PNPLA3I148M in mouse liver causes hepatic steatosis
John Zhong Li, … , Jonathan C. Cohen, Helen H. Hobbs
John Zhong Li, … , Jonathan C. Cohen, Helen H. Hobbs
Published October 1, 2012
Citation Information: J Clin Invest. 2012;122(11):4130-4144. https://doi.org/10.1172/JCI65179.
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Research Article Hepatology

Chronic overexpression of PNPLA3I148M in mouse liver causes hepatic steatosis

Abstract

A genetic variant in PNPLA3 (PNPLA3I148M), a triacylglycerol (TAG) hydrolase, is a major risk factor for nonalcoholic fatty liver disease (NAFLD); however, the mechanism underlying this association is not known. To develop an animal model of PNPLA3-induced fatty liver disease, we generated transgenic mice that overexpress similar amounts of wild-type PNPLA3 (PNPLA3WT) or mutant PNPLA3 (PNPLA3I148M) either in liver or adipose tissue. Overexpression of the transgenes in adipose tissue did not affect liver fat content. Expression of PNPLA3I148M, but not PNPLA3WT, in liver recapitulated the fatty liver phenotype as well as other metabolic features associated with this allele in humans. Metabolic studies provided evidence for 3 distinct alterations in hepatic TAG metabolism in PNPLA3I148M transgenic mice: increased formation of fatty acids and TAG, impaired hydrolysis of TAG, and relative depletion of TAG long-chain polyunsaturated fatty acids. These findings suggest that PNPLA3 plays a role in remodeling TAG in lipid droplets, as they accumulate in response to food intake, and that the increase in hepatic TAG levels associated with the I148M substitution results from multiple changes in hepatic TAG metabolism. The development of an animal model that recapitulates the metabolic phenotype of the allele in humans provides a new platform in which to elucidate the role of PNLPA3I148M in NAFLD.

Authors

John Zhong Li, Yongcheng Huang, Ruchan Karaman, Pavlina T. Ivanova, H. Alex Brown, Thomas Roddy, Jose Castro-Perez, Jonathan C. Cohen, Helen H. Hobbs

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Figure 4

Hepatic lipid synthesis in vivo or in primary hepatocytes from nontransgenic and PNPLA3I148M transgenic mice.

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Hepatic lipid synthesis in vivo or in primary hepatocytes from nontransg...
(A) Incorporation of 3H-glycerol into TAG. Tissue was collected from 8-week-old male mice (n = 4/group) 30 minutes after intraperitoneal injection with 3H-glycerol (1.6 nmol/mouse). (B–D) Incorporation of metabolic precursors into TAG in primary hepatocytes. Primary hepatocytes from mice of the indicated genotypes were isolated, attached to collagen-coated 6-well plates for 2 hours in triplicate, and then incubated with (B) 1.5 mM 14C-glycerol, (C) 1.0 mM 14C-acetate, or (D) 0.6 mM 14C-oleic acid for the indicated times. Lipids were extracted from the cells, and TAG was isolated by TLC as described in the Methods. (E) Measurement of LPAAT activity in membranes and lipid droplets isolated from mice of the indicated genotypes (n = 4/group), as described in the Methods. For the lipid droplet fraction, a total of 2 mg protein was added to 200 ml of buffer (50 mM TrisCl, pH 7.4) plus 200 mM LPA and 5.5 mM 14C-oleoyl-CoA. For the membrane fraction, an additional 50 mM oleoyl-CoA was added to the assay. The experiments were all repeated at least twice, and the results were similar.