Abstract
Although liver fatty acid-binding protein (L-FABP) is an important binding site for various hydrophobic ligands in hepatocytes, its in vivo significance is not understood. We have therefore created L-FABP null mice and report here their initial analysis, focusing on the impact of this mutation on hepatic fatty acid binding capacity, lipid composition, and expression of other lipid-binding proteins. Gel-filtered cytosol from L-FABP null liver lacked the main fatty acid binding peak in the fraction that normally comprises both L-FABP and sterol carrier protein-2 (SCP-2). The binding capacity for cis-parinaric acid was decreased >80% in this region. Molar ratios of cholesterol/cholesterol ester, cholesteryl ester/triglyceride, and cholesterol/phospholipid were 2- to 3-fold greater, reflecting up to 3-fold absolute increases in specific lipid classes in the order cholesterol > cholesterol esters > phospholipids. In contrast, the liver pool sizes of nonesterified fatty acids and triglycerides were not altered. However, hepatic deposition of a bolus of intravenously injected [14C]oleate was markedly reduced, showing altered lipid pool turnover. An increase of approximately 75% of soluble SCP-2 but little or no change of other soluble (glutathione S-transferase, albumin) and membrane (fatty acid transport protein, CD36, aspartate aminotransferase, caveolin) fatty acid transporters was measured. These results (i) provide for the first time a quantitative assessment of the contribution of L-FABP to cytosolic fatty acid binding capacity, (ii) establish L-FABP as an important determinant of hepatic lipid composition and turnover, and (iii) suggest that SCP-2 contributes to the accumulation of cholesterol in L-FABP null liver.
Highlights
Liver fatty acid-binding protein (L-FABP) is an important binding site for various hydrophobic ligands in hepatocytes, its in vivo significance is not understood
Given that liver fatty acid-binding protein (L-FABP) is expressed at very high levels (2–5% of cytosolic protein) in the differentiated hepatocyte [4, 5] and that these levels correlate well with lipid metabolism [2], it can be speculated that L-FABP contributes considerably to hepatic lipidbinding and lipid metabolism
The results showed that L-FABP was present at 18.1 Ϯ 3.0 ng/g in homogenates and at 45.2 Ϯ 3.3 ng/g in 105,000 ϫ g supernatants prepared from wild type livers
Summary
L-FABP, liver fatty acid-binding protein; SCP-2, sterol carrier protein-2; SCP-x, sterol carrier protein-x/3ketoacyl-CoA thiolase; FATP, fatty acid transport protein; FAT/CD36, fatty acid translocase; PBS, phosphate-buffered saline; GST, glutathione S-transferase. The only firmly established function of FABPs is the reversible binding of hydrophobic ligands, and these proteins do not exhibit any enzymatic function or energy requirement This suggests that these proteins play passive (facilitative) roles that, almost by definition, are strongly dependent on the cellular context. An in vivo approach is probably needed to elucidate the physiologically relevant roles of this protein Within this context, a deletional approach is likely to be more revealing than an overexpression approach, because L-FABP is extremely highly expressed even under basal conditions. We have decided to create L-FABP null mice by homologous recombination in embryonic stem cells, and the present paper reports their initial analysis We have focused this analysis on the impact of this mutation on lipid composition, lipid binding, and the expression of other known fatty acid-binding proteins
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