Peroxisomal and Mitochondrial Oxidation of Fatty Acids in the Heart, Assessed from the 13C Labeling of Malonyl-CoA and the Acetyl Moiety of Citrate

Abstract

We previously showed that a fraction of the acetyls used to synthesize malonyl-CoA in rat heart derives from partial peroxisomal oxidation of very long and long-chain fatty acids. The 13 C labeling ratio (malonylCoA)/(acetyl moiety of citrate) was >1.0 with 13 C-fatty acids, which yields [13 C]acetyl-CoA in both mitochondria and peroxisomes and < 1.0 with substrates, which yields [13 C]acetyl-CoA only in mitochondria. In this study, we tested the influence of 13 C-fatty acid concentration and chain length on the labeling of acetyl-CoA formed in mitochondria and/or peroxisomes. ]acetate, the labeling ratio was <0.7 at all concentrations. Therefore, in rat heart, (i) n-fatty acids shorter than 8 carbons do not undergo peroxisomal oxidation, (ii) octanoate undergoes only one cycle of peroxisomal ␤-oxidation, (iii) there is no detectable transfer to the mitochondria of acetyl-CoA from the cytosol or the peroxisomes, and (iv) the capacity of C 2 -C 18 fatty acids to generate mitochondrial acetyl-CoA decreases with chain length.Malonyl-CoA plays an important role in the regulation of fuel selection by the heart through its potent inhibition of carnitine palmitoyltransferase-I and of mitochondrial fatty acid oxidation (1). Malonyl-CoA is synthesized by acetyl-CoA carboxylase 2, which appears to be attached to the outer membrane of mitochondria facing the cytosol (2, 3). Because the heart is a non-lipogenic organ, the only known fate of malonyl-CoA is its decarboxylation by malonyl-CoA decarboxylase. Thus acetylCoA carboxylase and malonyl-CoA decarboxylase catalyze an acetyl-CoA 3 malonyl-CoA 3 acetyl-CoA substrate cycle that regulates malonyl-CoA concentration and mitochondrial fatty acid oxidation. The regulation of acetyl-CoA carboxylase and malonyl-CoA decarboxylase in the heart has been extensively investigated. For recent reviews, see Refs. 4 -8.Because the cytosolic concentrations of acetyl-CoA and malonyl-CoA are much lower than the K m of acetyl-CoA carboxylase and malonyl-CoA decarboxylase for their corresponding substrates, the concentrations of these substrates may regulate the fluxes through the enzymes. We recently measured the turnover of malonyl-CoA in isolated rat hearts perfused with NaH 13 CO 3 (9). Under conditions where the malonyl-CoA concentration varied over a 5-fold range, this concentration was directly proportional to the acetyl-CoA carboxylase flux, but the total activities of acetyl-CoA carboxylase and malonyl-CoA decarboxylase (measured in tissue extracts) did not vary. This strongly supports the notion that in the heart the cytosolic concentration of acetyl-CoA is a major determinant of the concentration of malonyl-CoA and of the rate of mitochondrial fatty acid oxidation.The acetyl-CoA used by cytosolic acetyl-CoA carboxylase is believed to be transferred from mitochondria either via acetylcarnitine or citrate and ATP-citrate lyase. We recently showed that a fraction of this acetyl-CoA is derived from the partial peroxisomal oxidation of long-chain or very long-chain fatty acids (10). Thi...

Probing peroxisomal β-oxidation and the labelling of acetyl-CoA proxies with [1-13C]octanoate and [3-13C]octanoate in the perfused rat liver

“…The labelling of the C1 + 2 moiety of BHB does not reflect that of the acetyl-CoA that enters the citric acid cycle. Our data also clearly demonstrate that octanoate can undergo two cycles of peroxisomal β-oxidation in liver, compared with one cycle in heart [31]. The relative labelling of the acetyl moiety of citrate and free acetate allows us to probe peroxisomal fatty acid oxidation in liver.…”

“…This dilution results from the formation of unlabelled acetyl-CoA from the peroxisomal β-oxidation of unlabelled endogenous long-chain fatty acids. It thus appears that, although octanoate can undergo two cycles of peroxisomal β-oxidation in liver (compared with one cycle in heart [31]), a greater fraction of the first acetyl of octanoate undergoes peroxisomal β-oxidation compared with the second acetyl.…”

Direct noninvasive estimation of myocardial tricarboxylic acid cycle flux in vivo using hyperpolarized 13C magnetic resonance

“…We showed that it is possible to estimate myocardial V TCA in vivo through the time-resolved 13 C MRS measurement of [5][6][7][8][9][10][11][12][13] C]citrate following the injection of hyperpolarized [1-13 C]acetate. V TCA and the dynamics of 13 C citrate labeling were independent on the amount of injected substrate and did not correlate with the dynamics of 13 C acetylcarnitine labeling.…”

“…The signal decay of precursor and metabolites results from a combination of the effects of longitudinal relaxation (characterized by the time constant T 1 ), RF excitation, and biochemical conversion and all time courses were corrected for the effect of repeated RF excitations. The T 1 of [1-13 C]acetylcarnitine was set to 14.9 s, as previously determined in vivo [35], and that of [5][6][7][8][9][10][11][12][13] C]citrate to 20 s [46]. The acetate signal decay was treated as free parameter.…”

“…The reported absence of CAT from the cytosol, may suggest two metabolic routes of acetate, (i) passing by the peroxisomal centers to enter the mitochondria as acetylcarnitine, or (ii) a direct entry via mitochondrial ACS. There have been conflicting reports on the involvement of CAT and acetylcarnitine in the transfer of acetyl groups from peroxisomes to the cytosol as discussed in [11], and therefore the conversion of acetate to acetylCoA via mitochondrial ACS seems most likely (Fig. 1).…”