Although liver fatty acid-binding protein (L-FABP) isan 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 family (1-3), is found in the liver, intestine, and kidney, but only in liver is it not co-expressed with other members of its family. L-FABP is known to bind fatty acids and various other hydrophobic molecules, although its actual contribution to the lipid-binding capacity of liver cytosol is not known. Given that 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. Work with cell-free systems and transfected cells has further strengthened this view. For example, in cell-free preparations L-FABP was shown to stimulate the esterification of oleic acid while inhibiting that of palmitic acid (6). L cells overexpressing L-FABP show increased rates of fatty acid uptake and esterification (7) as well as increased contents of phospholipid and cholesterol esters (8, 9). HepG2 hepatoma cells expressing an L-FABP antisense RNA showed a dose-dependent reduction of fatty acid uptake (10). Furthermore, overexpression of L-FABP in McA-RH7777 hepatoma cells incubated with palmitic acid decreased the synthesis and secretion of triglycerides while increasing beta oxidation and the secretion of apolipoprotein B100 (11). Thus, the various in vitro systems have allowed researchers to propose specific functions of L-FABP in vivo.However, in vitro studies of FABPs have inherent limitations. 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. One context of the highly expressed L-FABP is the highly differentiated hepatocyte, a cell type featuring an intense lipid metabolism that is not eas...

Liver fatty acid binding protein (L-FABP) has been proposed to limit the availability of long-chain fatty acids (LCFA) for oxidation and for peroxisome proliferator-activated receptor alpha (PPAR-alpha), a fatty acid binding transcription factor that determines the capacity of hepatic fatty acid oxidation. Here, we used L-FABP null mice to test this hypothesis. Under fasting conditions, this mutation reduced beta-hydroxybutyrate (BHB) plasma levels as well as BHB release and palmitic acid oxidation by isolated hepatocytes. However, the capacity for ketogenesis was not reduced: BHB plasma levels were restored by octanoate injection; BHB production and palmitic acid oxidation were normal in liver homogenates; and hepatic expression of key PPAR-alpha target (MCAD, mitochondrial HMG CoA synthase, ACO, CYP4A3) and other (CPT1, LCAD) genes of mitochondrial and extramitochondrial LCFA oxidation and ketogenesis remained at wild-type levels. During standard diet, mitochondrial HMG CoA synthase mRNA was selectively reduced in L-FABP null liver. These results suggest that under fasting conditions, hepatic L-FABP contributes to hepatic LCFA oxidation and ketogenesis by a nontranscriptional mechanism, whereas L-FABP can activate ketogenic gene expression in fed mice. Thus, the mechanisms whereby L-FABP affects fatty acid oxidation may vary with physiological condition.

BackgroundThe fetal cortical neuroepithelium is a mosaic of distinct progenitor populations that elaborate diverse cellular fates. Ethanol induces apoptosis and interferes with the survival of differentiating neurons. However, we know little about ethanol's effects on neuronal progenitors. We therefore exposed neurosphere cultures from fetal rat cerebral cortex, to varying ethanol concentrations, to examine the impact of ethanol on stem cell fate.ResultsEthanol promoted cell cycle progression, increased neurosphere number and increased diversity in neurosphere size, without inducing apoptosis. Unlike controls, dissociated cortical progenitors exposed to ethanol exhibited morphological evidence for asymmetric cell division, and cells derived from ethanol pre-treated neurospheres exhibited decreased proliferation capacity. Ethanol significantly reduced the numbers of cells expressing the stem cell markers CD117, CD133, Sca-1 and ABCG2, without decreasing nestin expression. Furthermore, ethanol-induced neurosphere proliferation was not accompanied by a commensurate increase in telomerase activity. Finally, cells derived from ethanol-pretreated neurospheres exhibited decreased differentiation in response to retinoic acid.ConclusionThe reduction in stem cell number along with a transient ethanol-driven increase in cell proliferation, suggests that ethanol promotes stem to blast cell maturation, ultimately depleting the reserve proliferation capacity of neuroepithelial cells. However, the lack of a concomitant change in telomerase activity suggests that neuroepithelial maturation is accompanied by an increased potential for genomic instability. Finally, the cellular phenotype that emerges from ethanol pre-treated, stem cell depleted neurospheres is refractory to additional differentiation stimuli, suggesting that ethanol exposure ablates or delays subsequent neuronal differentiation.

Embryonic cerebral cortical neuroepithelial-derived precursors secrete high levels of several angiogenic and neural-growth-promoting cytokines as they differentiate into neurons. Our data collectively suggest that ethanol exposure during the period of neuroepithelial proliferation significantly disrupts cytokine signals that are required for the support of emerging neurovascular networks, and the maintenance of neural stem cell beds.