OBJECTIVE: Heart failure is often preceded by distinct metabolic alterations. However, this shift cannot be sustained indefinitely and typically leads to cardiac dysfunction and arrest. These alterations are thought to be caused by mitochondrial dysfunction leading to the accumulation of acyl-CoAs. To compensate, the injured heart shifts from oxidation of long chain fatty acids (C16) to metabolism of substrates that can replenish the TCA cycle. This is because these fatty acids are restricted to catabolism to acetyl-CoA, for which the enzymes are saturated. By contrast, odd chain fatty acids are metabolized to propionate. The enzymes specific for propionate are freely available for metabolism to succinate leading to TCA cycle anaplerosis. Propionate conversion to propionyl-CoA is the first and major step for its sequestration and metabolism. Moreover, the mitochondrial acyl-CoA synthetase (ACSS) responsible for this conversion is highly expressed in the heart. Hence, we hypothesize that 2-[18 F]-fluoropropionate ([18 F]-FPA) would significantly accumulate in the failing heart and can be imaged by positron emission tomography (PET). We addressed our hypothesis by studying the substrate behavior of FPA for ACSS, and by metabolic profiling of HepG2 cells incubated with FPA. We also successfully imaged heart uptake of [18 F]-FPA in healthy rats by PET. METHODS: The ability of acetate, propionate, and FPA to act as ACSS substrates was determined by a colorimetric pyrophosphate assay. Metabolites were determined by LC/MS in lysates of HepG2 cells incubated with basal media, 10 mM propionate or 10 mM FPA. Racemic 18 F-FPA was synthesized from ethyl-2-bromopropionate by standard nucleophilic substitution. For PET, Sprague-Dawley rats were injected with 250-500 µCi (9.25-18.5 MBq) of [18 F]-FPA, and a 60-minute dynamic PET scan was immediately acquired. RESULTS: The kinetic characteristics for the reaction of ACSS acting on acetate (Km = 4.94 mM, Vmax = 0.25 nmols/sec), propionate (Km = 24.20 mM, Vmax = 0.19 nmols/sec), and 2-fluoropropionoate (V = 0.05 nmols/sec) were determined. We also observed a reduction of free CoA in HepG2 cells incubated in 10 mM propionate (4.4 ± 1.2-fold) or 10 mM FPA (3.09 ± 0.96-fold). The kinetics of 18 F-FPA uptake into the healthy heart was assessed by PET and determined to be maximal at 10 minutes post-injection. CONCLUSIONS: We identified [18 F]-FPA, and potentially other odd chain fatty acids, as promising metabolic tracers for imaging anaplerosis in the injured heart. FPA effectively decreases free CoA in HepG2 cells, which indicates the formation of acyl-CoAs. Moreover, FPA exhibits some substrate activity for ACSS, representing a major conduit by which it is metabolized in the heart. The poor substrate activity of FPA is ideal as it precludes metabolism by the highly active and specific ACSS enzymes in the liver. Albeit we expect FPA to accumulate in the heart due to its high ACSS expression which is induced in energy deficient states that are characteristic of heart failure. In support of this idea, we demonstrated that [18 F]-FPA is taken up by the healthy heart. We expect the metabolic alterations resulting from ensuing heart failure are sufficient to increase the uptake of [18 F]-FPA in these tissues. Our long-term aim is to develop 18 F-labeled short chain fatty acids, such as [18 F]-FPA, that can be used to diagnose at-risk patients. The early detection of heart failure by non-invasive means is critical for the development of interventions to reduce the severity and aid recovery from cardiac disease.
|Original language||English (US)|
|Journal||FASEB journal : official publication of the Federation of American Societies for Experimental Biology|
|State||Published - May 1 2022|
ASJC Scopus subject areas
- Molecular Biology