Role of CD36 in membrane transport and utilization of long-chain fatty acids by different tissues.

The transmembrane glycoprotein CD36 has been identified in isolated cell studies as a putative transporter of long-chain fatty acids. To examine the physiological role of CD36, we studied FA uptake and metabolism by tissues of CD36 null mice after injection with two fatty acid analogs. Compared to controls, uptake was substantially reduced (50-80%) in heart, skeletal muscle, and adipose tissues of null mice. The reduction in uptake was associated with a large decrease in fatty acid incorporation into triglycerides, which could be accounted for by an accumulation of diacylglycerides. Thus CD36 facilitates a major fraction of fatty acid uptake by myocardial, skeletal muscle, and adipose tissues, where it is highly expressed. Its role in other tissues where its expression is low and cell-specific could not be determined in these studies.

Defective fatty acid uptake in the spontaneously hypertensive rat is a primary determinant of altered glucose metabolism, hyperinsulinemia, and myocardial hypertrophy.

Genetic linkage studies implicated deficiency of CD36, a membrane fatty acid (FA) transporter, in the hypertriglyceridemia and hyperinsulinemia of the spontaneously hypertensive rat (SHR). In this study we determined whether loss of CD36 function in FA uptake is a primary determinant of the SHR phenotype. In vivo, tissue distribution of iodinated, poorly oxidized beta-methyliodophenyl pentadecanoic acid (BMIPP) was examined 2 h after its intravenous injection. Fatty acid transport was also measured in vitro over 20 to 120 s in isolated adipocytes and cardiomyocytes obtained from SHR and from a congenic line (SHRchr4) that incorporates a piece of chromosome 4 containing wild-type CD36. SHR heart and adipose tissue exhibited defects in FA uptake and in conversion of diglycerides to triglycerides that are similar to those observed in the CD36 null mouse. However, a key difference in SHR tissues is that fatty acid oxidation is much more severely impaired than fatty acid esterification, which may underlie the 4-5-fold accumulation of free BMIPP measured in SHR muscle. Studies with isolated adipocytes and cardiomyocytes directly confirmed both the defect in FA transport and the fact that it is underestimated by BMIPP. Heart, oxidative muscle, and adipose tissue in the SHR exhibited a large increase in glucose uptake measured in vivo using [(18)F]fluorodeoxyglucose. Supplementation of the diet with short-chain fatty acids, which do not require CD36-facilitated transport, eliminated the increase in glucose uptake, the hyperinsulinemia, and the heart hypertrophy in the SHR. This indicated that lack of metabolic energy consequent to deficient FA uptake is the primary defect responsible for these abnormalities. Hypertension was not alleviated by the supplemented diet suggesting it is unrelated to fuel supply and any contribution of CD36 deficiency to this trait may be more complex to determine. It may be worth exploring whether short-chain FA supplementation can reverse some of the deleterious effects of CD36 deficiency in humans, which may include hypertrophic cardiomyopathy.

Defective uptake and utilization of long chain fatty acids in muscle and adipose tissues of CD36 knockout mice.

The transmembrane protein CD36 has been identified in isolated cell studies as a putative transporter of long chain fatty acids. In humans, an association between CD36 deficiency and defective myocardial uptake of the fatty acid analog 15-(p-iodophenyl)-3-(R, S)-methyl pentadecanoic acid (BMIPP) has been reported. To determine whether this association represents a causal link and to assess the physiological role of CD36, we compared tissue uptake and metabolism of two iodinated fatty acid analogs BMIPP and 15-(p-iodophenyl) pentadecanoic acid (IPPA) in CD36 null and wild type mice. We also investigated the uptake and lipid incorporation of palmitate by adipocytes isolated from both groups. Compared with wild type, uptake of BMIPP and IPPA was reduced in heart (50-80%), skeletal muscle (40-75%), and adipose tissues (60-70%) of null mice. The reduction was associated with a 50-68% decrease in label incorporation into triglycerides and in 2-3-fold accumulation of label in diglycerides. Identical results were obtained from studies of [(3)H]palmitate uptake in isolated adipocytes. The block in diglyceride to triglyceride conversion could not be explained by changes in specific activities of the key enzymes long chain acyl-CoA synthetase and diacylglycerol acyltransferase, which were similar in tissues from wild type and null mice. It is concluded that CD36 facilitates a large fraction of fatty acid uptake by heart, skeletal muscle, and adipose tissues and that CD36 deficiency in humans is the cause of the reported defect in myocardial BMIPP uptake. In CD36-expressing tissues, uptake regulates fatty acid esterification at the level of diacylglycerol acyltransferase by determining fatty acyl-CoA supply. The membrane transport step may represent an important control site for fatty acid metabolism in vivo.

Reversible binding of long-chain fatty acids to purified FAT, the adipose CD36 homolog.

Transport of long-chain fatty acids into rat adipocytes was previously shown to be inhibited by the reactive derivative sulfosuccinimidyl oleate consequent to its binding to a membrane protein FAT, which is homologous to CD36. In this report, the ability of the purified protein to bind native fatty acids was investigated. CD36 was isolated from rat adipocytes by phase partitioning into Triton X-114 followed by chromatography on DEAE and then on wheat germ agglutinin. Fatty acid binding was determined by incubating CD36, solubilized in buffer containing 0.1 Triton X-100, with fatty acids at 37 degrees C, and then by adsorbing the unbound ligand with Lipidex 1,000 at 0 degrees C. Bovine serum albumin was used as a positive control and gelatin, a protein that does not bind fatty acids, as a negative control. Measurements with albumin yielded reproducible binding values which were not altered by the presence of 0.1% Triton X-100. Under the same conditions, gelatin yielded reproducibly negative measurements that did not differ significantly from zero. CD36 bound various long-chain fatty acids at low ligand to protein ratios. Warming the protein-FA-Lipidex mixture to 37 degrees C removed the FA off the protein. Thus, binding was reversible and distinct from the palmitoylation of the protein known to occur on an extracellular domain. Comparison of the predicted secondary sequence of CD36 with that of human muscle fatty acid binding protein suggested that a potential binding site for the fatty acid on CD36 may exist in its extracellular segment between residues 127 and 279.